Pattern Of Chromosomal Aberrations Research Articles

Quantitative Chemical Proteomics Reveals New Potential Drug Targets in Head and Neck Cancer

Tumors of the head and neck represent a molecularly diverse set of human cancers, but relatively few proteins have actually been shown to drive the disease at the molecular level. To identify new targets for individualized diagnosis or therapeutic intervention, we performed a kinase centric chemical proteomics screen and quantified 146 kinases across 34 head and neck squamous cell carcinoma (HNSCC) cell lines using intensity-based label-free mass spectrometry. Statistical analysis of the profiles revealed significant intercell line differences for 42 kinases (p < 0.05), and loss of function experiments using siRNA in high and low expressing cell lines identified kinases including EGFR, NEK9, LYN, JAK1, WEE1, and EPHA2 involved in cell survival and proliferation. EGFR inhibition by the small molecule inhibitors lapatinib, gefitinib, and erlotinib as well as siRNA led to strong reduction of viability in high but not low expressing lines, confirming EGFR as a drug target in 10-20% of HNSCC cell lines. Similarly, high, but not low EPHA2-expressing cells showed strongly reduced viability concomitant with down-regulation of AKT and ERK signaling following EPHA2 siRNA treatment or EPHA1-Fc ligand exposure, suggesting that EPHA2 is a novel drug target in HNSCC. This notion is underscored by immunohistochemical analyses showing that high EPHA2 expression is detected in a subset of HNSCC tissues and is associated with poor prognosis. Given that the approved pan-SRC family kinase inhibitor dasatinib is also a very potent inhibitor of EPHA2, our findings may lead to new therapeutic options for HNSCC patients. Importantly, the strategy employed in this study is generic and therefore also of more general utility for the identification of novel drug targets and molecular pathway markers in tumors. This may ultimately lead to a more rational approach to individualized cancer diagnosis and therapy.

O56. Genome wide profiling of tongue and cheek cancer using high resolution array based CGH

Introduction: Tongue and cheek cancer have different behaviors. In order to understand these behaviors, there is a need to look into the chromosomal alterations and gene pathways that maybe associated with oral cancer at these sites. Therefore, the objective of this study is to determine the differences in chromosomal aberrations and gene pathways involved in tongue and cheek cancer using high resolution array CGH. Methods: A genome wide screening with array CGH (SurePrint G3 CGH 1x1M microarray) was performed using gDNA from 20 snap frozen fresh tissues consisting of 12 tongue and 8 cheek oral squamous cell carcinomas (samples from the Malaysian Oral Cancer Data and Tumour Bank System [MOCDTBS] coordinated by OCRCC-UM). Cytosure Software was used to detect the chromosomal aberrations and candidate genes related to the selected regions. Pathway analysis was done using MetaCore™software for selected genes. Results: The mean number of chromosomal aberrations per tumour for tongue cancer (22 ± 24.97) was higher than cheek cancer (8.38 ± 11.98). The most common amplified regions in tongue cancers were 8q24.22 (33.33%), 8q24.3 (33.33%), 11q13.1 (33.33%), 11q13.2 (33.33%), 12q13.13 (33.33%), 14q32.33 (33.33%) and for cheek cancer the most common amplified region was 22q12.3 (25%). For the deleted regions, the most common for tongue cancer were 2q21.1 (16.67%), 6q21 (16.67%) and for cheek cancer were 2q22.1 (25%), 7q35 (25%), 19q13.33 (25%). The most significant pathway based on p < 0.001 involved in tongue cancer is cell adhesion ECM remodeling pathway Among the genes involved in amplified region, were EGFR, MMP1, MMP10, MMP12, MMP3, MMP7 and MMP9. For cheek cancer, the cancer associated significant pathway was apoptosis and survival (the role of CDK5 in neuronal death and survival). An oncogene located at 8p12 was identified in this pathway is NRG1. Discussion: This study showed a different pattern of chromosomal aberrations in tongue and cheek cancer with different significant gene pathways. Tongue cancer behaves more aggressively than cheek cancer which may be due to the involvement of ECM remodeling pathway, where MMP family which are the proteolytic enzyme that degrade various component in the extracellular matrix would favor invasion and metastasis.

Activation of common fragile sites as a possible cause of chromosome 8 instability in colorectal cancer

Colorectal cancer (CRC) ranks among the leading causes of cancer death worldwide. The identification of genes that influence colorectal cancer susceptibility and progression therefore remains an essential goal of basic and clinical research. The majority of CRC tumours are microsatellite stable and characteristically show chromosomal instability with losses of large chromosomal regions. Specific patterns of structural chromosomal aberrations and oncogene amplification have been associated with this tumour type.

Use of Immunohistochemistry in the Diagnosis of Melanocytic Lesions: Applications and Pitfalls

The accurate diagnosis of melanocytic lesions is essential for the adequate clinical management of the patients. Besides the histopathologic examination, immunohistochemical studies are often used as an adjunct in distinguishing melanocytic lesions from tumors with different origin or between benign and malignant melanocytic lesions. In the first part of this article, we analyze data on currently used immunohistochemical markers, with special emphasis on their applicability to clinical practice, and underline their potential pitfalls. The pathogenesis of malignant transformation of melanocytes is not completely understood. Recent studies report that various melanoma progression markers are preferentially expressed in benign or malignant melanocytic lesions or show different expression in subsequent stages of tumor development. Furthermore, in recent years, emerging genetic studies suggest that there are distinctive patterns of chromosomal aberrations in different subtypes of melanoma that can be altered by newly developed targeted therapies. In the second part of our article, we will discuss the most significant progression markers in melanoma that can be detected by immunohistochemistry and their potential usefulness for diagnosis, prognosis, staging or as therapeutic targets.

Abstract 2158: Loss of heterozygosity at chromosome 3p in ductal and lobular breast carcinomas

Abstract The two main histological types of infiltrating breast cancer, lobular (ILC) and the more common ductal (IDC) carcinoma are morphologically and clinically distinct revealing different patterns of chromosomal aberrations and gene expression. Loss of heterozygosity (LOH) at chromosome 3p is frequently observed in breast tumors and a whole genome study utilizing a SNP array assay found a significantly higher frequency of LOH on 3p in ductal than lobular tumors. In order to investigate the molecular alterations at 3p associated with these breast cancer subtypes we analyzed LOH in 118 IDC and 18 ILC samples using seven microsatellite markers. Genomic DNA was obtained from frozen tumor tissue and peripheral blood was used as a normal reference. All samples were genotyped on a MegaBace® system and analyzed using Fragment Profiler® Version 2.1 software. Significance was determined using Chi-square test (X2). A significant higher frequency of LOH was observed in IDC (62.7%) when compared with ILC (38.9%) (X2=4.60 P&amp;lt;0.05). When the markers were analyzed separately, LOH at three of them, D3S1307 in 3p26.3, D3S1286 in 3p24.3 and D3S1300 (intragenic FHIT) in 3p14.2, were significantly more frequent in ductal than lobular tumors (X2= 4.66 P&amp;lt;0.05; X2= 3.99 P&amp;lt;0.05 and X2= 5.43 P&amp;lt;0.025, respectively). D3S1307 marker showed the highest frequency of LOH in IDC (47.8%) and associations between loss of this marker and loss of estrogen and progesterone receptors were found in these samples (X2= 4.61 P&amp;lt;0.05 and X2= 4.49 P&amp;lt;0.05, respectively) suggesting that there are genes on this region involved in the control of differentiation of ductal tumors. For lobular tumors, the highest loss (53.3%) was seen for D3S1274 on 3p12.3 (intragenic DUTT1). In addition to confirm previous studies these findings suggest that specific losses on 3p (3p26.3, 3p24.3 and 3p14.2) are more common in IDC when compared with ILC breast tumors. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2158.

Abstract 1757: Patterns of clonal chromosomal aberrations in invasive breast cancer compared to adjacent DCIS lesions detected by a ten-probe FISH panel

Abstract Ductal carcinoma in situ (DCIS) refers to lesions with malignant epithelial cells that have not invaded the surrounding stroma. These lesions are confined to the ducts of the breast. It is thought that DCIS represents a stage that precedes invasive ductal carcinoma (IDC). However the linkage between these two lesions is still obscure. In order to analyze the patterns of genetic alterations as an approach to understand the correlation between DCIS and IDC, we developed four panels of FISH probes consisting of a total of eight gene specific probes and two centromeric probes and conducted a systematic FISH evaluation of breast tumor specimens containing both lesions. The gene probes tested for oncogenes and tumor suppressor genes specific for breast tumorigenesis while the centromeric probes were included to reflect and adjust for the ploidy of the cells. Objectives Determine gain and loss patterns of the oncogene and tumor suppressor gene probes within DCIS and IDC Compare similarities in the genetic make-up of DCIS and corresponding IDC lesions from the same case Characterize clonal evolution from DCIS lesions to IDC The database of the “Breast Susceptibility Study” conducted at the Naval Medical Center, Bethesda, Maryland was searched for patients whose pathology reports were positive for breast carcinoma with neighboring DCIS. Archived paraffin blocks were analyzed by a pathologist for the presence of invasive cancer and DCIS. Representative areas were marked on thick sections. The material was micro-dissected and disintegrated using the Hedley method. The resulting single cell suspensions were cytospun onto slides. The best cytospin for each lesion was. hybridized with four FISH probe panels subsequently. An Imaging relocation software allowed us to assess the same cells for all four sets of probes. For each lesion, manual counting of 150-200 nuclei was performed and tabulated into an excel spreadsheet to allow analysis of signal patterns. In collaboration with NCBI, algorithms were developed to compare clonal patterns. Preliminary data show that there are often major clones both in the DCIS and the IDC that present strong similarities to each other. However, the clones can differ in specific gains or losses of the oncogenes and tumor suppressor genes tested. The signals depicted in the table represent an attempt to delineate major clones within the DCIS and IDC populations. This study will improve our understanding of the molecular events that are involved in the progression from pre-invasive to invasive cancer and may lead to the development of prognostic biomarkers that can predict this transition. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1757.

Abstract 1493: High-resolution comparative genomic hybridization of preneoplastic and invasive esophageal squamous cell carcinoma: Identification of the earliest preneoplastic changes

Abstract Background: Esophageal cancer causes &amp;gt; 400,000 deaths/year in the world. Clinically useful early detection methods must identify these cancers and their precursor lesions at more curable stages. We conducted comparative genomic hybridization analyses of preneoplastic and invasive esophageal squamous cell carcinoma to look for gene copy number patterns that might be useful as early detection markers. The current effort focuses on aberrations associated with mild squamous dysplasia, the earliest histologic form of squamous preneoplasia. Methods: DNA was extracted from either biopsies or surgical resections from 122 esophageal tissues spanning a spectrum including normal mucosa, squamous preneoplasia, and invasive squamous cell carcinoma. High-resolution micro-array based comparative genomic hybridization (Agilent) was used to measure regions of chromosomal gain or loss with an emphasis on identifying changes associated with the transition from normal to mild dysplasia. Results: Chromosomal number aberrations were identified that involve several chromosomes, occur early in the neoplastic process, and progress with disease severity. Specific chromosomal number aberrations associated with mild squamous dysplasia, the earliest form of squamous preneoplasia, were uniquely identified. Conclusions: Comparative genomic hybridization identifies patterns of chromosomal number aberrations that may distinguish patients with and without high-grade squamous dysplasia or cancer. The current effort also identifies specific changes associated with mild squamous dysplasia, the earliest form of preneoplasia, and this may help elucidate mechanisms related to the initiation of carcinogenesis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1493.

Positioning of necrotic lobular intraepithelial neoplasias (LIN, grade 3) within the sequence of breast carcinoma progression

Lobular intraepithelial neoplasia Grade 3 (LIN3) is a recently recognized variant of intraepithelial lobular neoplasia (LIN) of the breast composed of either uniform, generally small cells with massive lobular distension, pleomorphic cells, signet-ring cells, or any cell type with necrosis. In contrast to classic forms of LIN, there is no consensus on therapeutic strategies for LIN3. In part this is due to the paucity of molecular data that could assist in defining the relationship of LIN3 to classic LIN and carcinomas. In this study we have employed array comparative genomic hybridization to determine the patterns of chromosomal aberrations in nine LIN3 lesions. By comparison to array CGH data of 13 classic LIN lesions, we demonstrate that classic LIN and LIN3 share several recurrent changes, in particular gains of 1q and losses of 16q. Both aberrations are known to appear early in tumorigenesis and to be associated with good prognosis. However, apart from this overlap, there were a number of karyotypic features that were observed exclusively in LIN3. Clearly, this lesion was characterized by a significantly higher number of DNA copy number changes (9 vs. 31 on average), a considerable complexity of chromosomal rearrangements with more than 16 breakpoints in one chromosome and overlapping high copy amplifications encompassing a number of known oncogenes. Our data suggest that, at the genetic level, LIN3 represents a highly advanced lesion with considerable resemblance to carcinomas and, therefore, might represent the transition state from an intraepithelial neoplasm to breast carcinoma.

Pattern of chromosomal aberrations in primary liver cancers identified by comparative genomic hybridization

Pattern of chromosomal aberrations in primary liver cancers identified by comparative genomic hybridization

Similar Patterns of Chromosome Abnormalities in CML Occur in Addition to the Philadelphia Translocation with or without Tyrosine Kinase Inhibitor Treatment.

Abstract 3270Poster Board III-1In chronic myeloid leukemia (CML), the progress from chronic phase (CP) to accelerated (AP)/blast phase (BP) is frequently accompanied by cytogenetic evolution within the Philadelphia (Ph+) positive clone. It was shown that additional chromosomal alterations follow non-random patterns with frequent occurrence of “major route” (e.g. +Ph, i(17q), +8) and “minor route” changes (e.g. -Y, -7 or +21). Corresponding to the pre-imatinib era, Ph+ clonal cytogenetic evolution was still identified to worsen prognosis in patients receiving imatinib. However, whether the introduction of TKIs changed the pattern of additional chromosome aberrations, was not studied so far. We here compared two subgroups: 1.) 245 CML patients treated with tyrosine kinase inhibitors (TKIs) with clonal evolution in the Ph+ clone investigated in the Munich Leukemia Laboratory (MLL) between 2005–2009. 2.) 500 CML cases published in the Mitelman Database (http://cgap.nci.nih.gov/Chromosomes/Mitelman) before the year 2000 (i.e. before the introduction of imatinib into the treatment of CML). The 245 patients from our cohort were selected for this study based on the occurrence of Ph+ clonal evolution at diagnosis or during the course of CML. Patients received imatinib or 2nd generation TKIs. First, analysis was performed for the Mitelman cohort and for the MLL cohort. Then, analysis was separated for those patients from the MLL cohort who showed Ph+ clonal evolution already at diagnosis of CML before start of TKIs (n=91), and for those patients who developed Ph+ cytogenetic alterations during TKIs (n=154). The patterns of chromosomal gains and losses were analyzed with the support of the CyDAS cytogenetic data analysis system (http://www.cydas.org/OnlineAnalysis/). All 4 cohorts showed comparable patterns of chromosomal gains/losses in addition to the Philadelphia translocation: Most frequent were +8, +Ph chromosome, +19, and i(17)(q10), and -Y. Less frequent were -7 and +21. Therefore, no difference between the patterns of abnormalities dating from the pre-imatinib era (data from the Mitelman database) or after the introduction of TKI (MLL cohort) was obvious. Also, there was no difference in cytogenetic patterns between patients who showed Ph+ clonal evolution at diagnosis of CML already and those who acquired them during TKI treatment. In conclusion, the patterns of cytogenetic alterations in the Ph+ clones in CML are similar as investigated in the pre-TKI and the TKI eras and therefore are independent, i.e. of the time point of analysis at diagnosis or during follow-up and also treatment of CML. Disclosures:Disclosures: Haferlach:MLL Munich Leukemia Laboratory: Equity Ownership. Schnittger:MLL Munich Leukemia Laboratory: Equity Ownership. Weiss:MLL Munich Leukemia Laboratory: Employment. Kern:MLL Munich Leukemia Laboratory: Equity Ownership. Haferlach:MLL Munich Leukemia Laboratory: Equity Ownership.

Specific chromosomal imbalances as detected by array CGH in ependymomas in association with tumor location, histological subtype and grade

Ependymomas are glial neoplasms originating from the wall of the ventricles or from the spinal canal. The significance of histopathological features in accurately predicting biological behavior is still debated. Moreover, key molecular events in the pathogenesis of ependymoma are yet to be defined. The main objective of the present study was to identify specific patterns of chromosomal aberrations that correlate with tumor location, histological subtype and grade. Forty-five ependymoma samples were analyzed by 1-megabase resolution array comparative genomic hybridization (CGH). Association between clinical or histopathological parameters and the genomic alterations identified was evaluated. The most frequently detected chromosome (chr) abnormalities were gain of chr 7, 9, 12 and 15 and loss of chr 22. Co-occurrence of those five alterations characterized spinal ependymomas (P = 0.01). Myxopapillary ependymomas displayed a specific genomic profile defined by concurrent gain of chr 5, 7, 9, 16 and 18 (P = 0.0007). Overall, the number of chromosomal abnormalities detected was inversely correlated with the malignancy grade. Gain of chr 1q correlated with intracranial high-grade tumors (P = 0.002). Loss of chr 6q was mainly observed in infratentorial (P = 0.02) and World Health Organization (WHO) grade III (P = 0.04) lesions. Chr 10q loss was associated with high-grade ependymomas (P = 0.01). The +7/+9/+12/+15/-22 genomic profile is significantly associated with WHO grade II spinal ependymomas, whereas the +5/+7/+9/+16/+18 genomic pattern is specific of myxopapillary ependymomas. Identification of specific genomic imbalances at a given tumor location suggests that ependymomas from different central nervous system (CNS) regions represent genetically distinct diseases. Detecting genomic alterations associated with aggressive biological behavior may help stratify patients in high- and low-risk disease.

Chromosomal copy number changes supporting the classification of lentiginous junctional melanoma of the elderly as a subtype of melanoma

Recently the term lentiginous melanoma of the elderly has been suggested for a pattern of melanocytic neoplasia characterized by frequent occurrence in elderly patients, broad lentiginous growth pattern, with poorly cohesive nesting, suprabasilar extension of melanocytes and moderate cytological atypia. However, there are limited reported cases with follow-up information to confirm the malignant nature of these neoplasms. Using fluorescence in situ hybridization (FISH) targeting chromosomal loci that are frequently found to have copy number changes in melanoma, we evaluated cases of lentiginous junctional melanoma of the elderly in order to compare with the frequencies and patterns of chromosomal aberrations identified in other subtypes of melanoma. Previous studies have shown that by using a FISH assay targeting 6p25, 6q23, 11q13 and CEP6 with previously validated criteria, one could discriminate benign nevi from melanoma with high sensitivity and specificity. In this study, 16 of 19 cases (84%) showed sufficient copy number changes in one of the targeted chromosomal loci to meet FISH criteria for melanoma. A total of 17 control cases of lentiginous junctional nevi tested negative for all criteria. These findings support the classification of this pattern of melanocytic neoplasia as a subtype of melanoma.

Finding exclusively deleted or amplified genomic areas in lung adenocarcinomas using a novel chromosomal pattern analysis

BackgroundGenomic copy number alteration (CNA) that are recurrent across multiple samples often harbor critical genes that can drive either the initiation or the progression of cancer disease. Up to now, most researchers investigating recurrent CNAs consider separately the marginal frequencies for copy gain or loss and select the areas of interest based on arbitrary cut-off thresholds of these frequencies. In practice, these analyses ignore the interdependencies between the propensity of being deleted or amplified for a clone. In this context, a joint analysis of the copy number changes across tumor samples may bring new insights about patterns of recurrent CNAs.MethodsWe propose to identify patterns of recurrent CNAs across tumor samples from high-resolution comparative genomic hybridization microarrays. Clustering is achieved by modeling the copy number state (loss, no-change, gain) as a multinomial distribution with probabilities parameterized through a latent class model leading to nine patterns of recurrent CNAs. This model gives us a powerful tool to identify clones with contrasting propensity of being deleted or amplified across tumor samples. We applied this model to a homogeneous series of 65 lung adenocarcinomas.ResultsOur latent class model analysis identified interesting patterns of chromosomal aberrations. Our results showed that about thirty percent of the genomic clones were classified either as "exclusively" deleted or amplified recurrent CNAs and could be considered as non random chromosomal events. Most of the known oncogenes or tumor suppressor genes associated with lung adenocarcinoma were located within these areas. We also describe genomic areas of potential interest and show that an increase of the frequency of amplification in these particular areas is significantly associated with poorer survival.ConclusionAnalyzing jointly deletions and amplifications through our latent class model analysis allows highlighting specific genomic areas with exclusively amplified or deleted recurrent CNAs which are good candidate for harboring oncogenes or tumor suppressor genes.

Pattern of chromosomal aberrations in primary liver cancers identified by comparative genomic hybridization

Little is known about the molecular cytogenetic changes in cholangiocarcinoma and combined hepatocellular-cholangiocarcinoma, and on the prognostic significance of chromosomal imbalances in hepatocellular carcinoma. Seventy-eight cases of primary liver cancer with available median follow-up of 16.5 months, including 49 hepatocellular carcinomas, 22 cholangiocarcinomas, and 7 combined hepatocellular-cholangiocarcinomas, were examined by comparative genomic hybridization. In hepatocellular carcinoma, the most frequent changes were +8q (54%), -8p (54%), and +1q (42%), followed by -6q (35%), -4q (33%), -13q (29%), -14q (25%), -16q (19%), -17p (19%), +17q (17%), and +20q (15%). In comparison, cholangiocarcinoma had more gains, losses, and breakpoints than hepatocellular carcinoma or combined hepatocellular-cholangiocarcinoma, specifically more frequently -6q (91%), -3p (68%), -9p (55%), -14q (55%), -13q (45%), +1q (41%), +7q (36%), +7p (32%), and +8q (32%). Combined losses at 6q and 3p appeared to be highly characteristic for cholangiocarcinoma. In contrast, combined hepatocellular-cholangiocarcinoma shared frequent +1q (71%), +8q (57%), and -8p (57%) with hepatocellular carcinoma, but a tendency for higher numbers of imbalances with cholangiocarcinoma. Overall, higher numbers of changes, breakpoints, or gains appeared to carry unfavorable prognostic value among hepatocellular carcinomas, with higher numbers of gains retaining prognostic value among R0-resected hepatocellular carcinomas. Cholangiocarcinoma is characterized by combined losses at 6q and 3p and a tendency for chromosomal instability. On the other hand, combined hepatocellular-cholangiocarcinoma may share similar chromosomal changes with both hepatocellular carcinoma and cholangiocarcinoma, as reflected by common hepatocellular carcinoma-like +8q, +1q, and -8p and a tendency for cholangiocarcinoma-like chromosomal instability. In hepatocellular carcinoma, higher number of gains may prove an adverse prognostic parameter.

Genetic classification of oral and oropharyngeal carcinomas identifies subgroups with a different prognosis.

The common risk factors for oral and oropharyngeal cancer are tobacco smoking and alcohol consumption, and recently the human papillomavirus (HPV) was shown to be involved in a subgroup. HPV-positive and -negative carcinomas can be distinguished on basis of their genetic profiles. Aim of this study was to investigate patterns of chromosomal aberrations of HPV-negative oral and oropharyngeal squamous cell carcinomas (OOSCC) in order to improve stratification of patients regarding outcome. Thirty-nine OOSCCs were classified on basis of their genetic pattern determined by array comparative genomic hybridization (aCGH). Resulting groups were related to patient and tumor characteristics using the Fisher’s exact test and in addition to survival with the Kaplan–Meier and log rank tests. Classification distinguished three groups, one characterized by hardly any chromosomal aberration (N = 8) and another by a relatively high level (N = 26), and one with a very high level (N = 5) of chromosomal aberrations. This classification was significantly (p = 0.003) associated with survival, with the best survival in the genetically ‘silent’ group and the worst survival in the most aberrant group. The silent profile was significantly (p < 0.05) associated with wild-type TP53, an absence of alcohol consumption and a female gender. These carcinomas were negative for microsatellite instability. This classification of OOSCC was confirmed in an independent set of 89 oral carcinomas. In conclusion, the discovery of these new classes of oral and oropharyngeal cancer with unique genetic and clinical characteristics has important consequences for future basic and clinical studies.

Genetic Classification of Oral and Oropharyngeal Carcinomas Identifies Subgroups with a Different Prognosis

The common risk factors for oral and oropharyngeal cancer are tobacco smoking and alcohol consumption, and recently the human papillomavirus (HPV) was shown to be involved in a subgroup. HPV-positive and -negative carcinomas can be distinguished on basis of their genetic profiles. Aim of this study was to investigate patterns of chromosomal aberrations of HPV-negative oral and oropharyngeal squamous cell carcinomas (OOSCC) in order to improve stratification of patients regarding outcome. Thirty-nine OOSCCs were classified on basis of their genetic pattern determined by array comparative genomic hybridization (aCGH). Resulting groups were related to patient and tumor characteristics using the Fisher’s exact test and in addition to survival with the Kaplan–Meier and log rank tests. Classification distinguished three groups, one characterized by hardly any chromosomal aberration (N = 8) and another by a relatively high level (N = 26), and one with a very high level (N = 5) of chromosomal aberrations. This classification was significantly (p = 0.003) associated with survival, with the best survival in the genetically ‘silent’ group and the worst survival in the most aberrant group. The silent profile was significantly (p &lt; 0.05) associated with wild-type TP53, an absence of alcohol consumption and a female gender. These carcinomas were negative for microsatellite instability. This classification of OOSCC was confirmed in an independent set of 89 oral carcinomas. In conclusion, the discovery of these new classes of oral and oropharyngeal cancer with unique genetic and clinical characteristics has important consequences for future basic and clinical studies.

Pattern of Chromosomal Aberrations and IgVH Mutation Status in Patients with Monoclonal B-Cell Lymphocytosis (MBL)

The separation between chronic lymphocytic leukemia (CLL) and monoclonal B-cell lymphocytosis (MBL) with CLL-phenotype by the presence of more than 5,000/μl lymphocytes in peripheral blood has been debated due to the rather continuous spectrum of clinical and biological features of both entities. Furthermore, this separation is influenced by non-malignant cells since it is based on the total lymphocyte count and not only on the count of cells with CLL-phenotype. To broaden the insights into the genetic background of MBL at the boarder to CLL we analyzed 188 cases with less than 5,000/μl CLL-phenotype cells for chromosomal aberrations by fluorescence in situ hybridization (FISH) using a standard panel of probes as well as for their IgVH mutation status. In addition, in 116 of these cases conventional chromosome analysis has been performed. Patient characteristic were (median/range): age, 66.0/27.5–86.6 years; WBC count, 10,600/1,700–22,660/μl; cells with CLL-phenotype 26/4-94% and 3,081/264–4,998/μl, respectively. The proportion of aberrant/analyzed cases for the respective chromosome aberrations as determined by FISH were: 11q-, 10/182 (5.5%); +12, 39/183 (21.3%); 6q-, 2/182 (1.1%); 13q-, 88/185 (47.6%); 17p-, 3/183 (1.6%); and t(11;14), 8/183 (4.4%). In 57/185 (30.8%) cases no aberration was found by FISH. A comparison to a previous series of 518 CLL cases revealed similar frequencies for +12, 6q-, and t(11;14) but higher frequencies in CLL for 11q-, 5.5 vs. 13.1%, p=0.004; 13q-, 47.6 vs. 57.8%, p=0.020; and 17p-, 1.6 vs. 8.1%, p=0.001. Within the present series, a relation between the respective chromosome aberration and the count of cells with CLL-phenotype was found for 6q- vs. no 6q-, 1,214 vs. 2,910/μl, p=0.079; 13q- vs. no 13q-, 3,285 vs. 2,486/μl, p<0.001; t(11;14) vs. no t(11;14), 2,025 vs. 2,920/μl, p=0.087 but not for 11q-, +12, and 17p- aberrations. IgVH mutation status has been analyzed in 172 cases and was found unmutated (IgVHunmut) in 44 (25.6%) und mutated (IgVHmut) in 111 (64.5%). No clonal IgVH rearrangement was detectable in 17 (9.9%). A relation to the count of cells with CLL-phenotype was found: IgVHmut vs. IgVHunmut, 3,071 vs. 2,629/μl, p=0.052. Conventional chromosome analysis has been performed in addition in 116 of these cases resulting in 111 (95.7%) evaluable cases. In general, results obtained by FISH were confirmed: 11q-, n=3 (1.8%); +12, n=21 (21.3%); 6q- 1 (0.9%); 13q-, n=39 (35.1%), and t(11;14), n=5 (4.5%). Additional aberrations were found and included other trisomies (n=13), other balanced translocations/inversions (n=13), 14q- (n=8), loss of sex chromosome (n=3), t(14;18) (n=1), and 2q- (n=1). Thus, a total of 39 chromosome aberrations have been found in 111 cases as assessed by conventional chromosome analysis which could not be detected by FISH using a standard panel of probes. In 30 cases (27%) a normal karyotype was found. These data indicate that the genetic background of MBL is highly related to the one of classical CLL as defined today although 11q-, 13q- and 17p- occur more frequently in CLL. Chromosome aberrations are present in clearly more than half of the cases and an unmutated IgVH status can be detected in a quarter of cases. Similar to CLL, conventional chromosome analysis reveals a substantial number of aberrations not detectable by FISH using a standard panel of probes. It is suggested to further analyze the genetic spectrum of MBL and early CLL cases and its clinical impact and to challenge based on these data the rather arbitrarily defined cut-off values like 5,000/μl lymphocytes.

About 17% of AML with NPM1 mutations Show a Specific Pattern of Chromosome Aberrations but These Cases Do Not Differ Prognostically from AML with NPM1 Mutations Carrying a Normal Karyotype

AML with mutated nucleophosmin gene (AML NPM1mut) usually carries a normal karyotype and will be suggested as a provisional entity in the new WHO classification. Thus far, the impact of chromosome aberrations in AML NPM1mut has not been evaluated in detail. Aim of this study was to determine the incidence and prognostic impact of clonal chromosome aberrations in NPM1mut. We further compared this pattern to additional aberrations in AML with recurrent genetic aberrations: t(8;21)(q22;q22), inv(16)(p13q22)/t(16;16)(p13;q22), t(15;17)(q22;q12) and 11q23-abnormalities leading to an MLL-rearrangement. In total 415 AML (de novo AML: 392, s-AML: 11, t-AML: 12) showing an NPM1 mutation were analyzed by chromosome banding analysis. 71 of these showed clonal chromosome aberrations (17.1%; de novo AML: 63 (16.1%), s-AML: 5 (45.5%), t-AML: 3 (25%); de novo AML vs. s-AML: p=0.024). Overall, 111 chromosome aberrations were observed. The most frequent abnormalities were +8 (n=30), −Y (n=10), +4 (n=9), del(9q) (n=5), +21 (n=4), −7 (n=3), +5 (n=2), +10 (n=2), +13 (n=2),+18 (n=2), del(12p) (n=2), del(20q) (n=2), other non-recurrent balanced aberrations (n=6), other non-recurrent unbalanced aberrations (n=32). For comparison 63 AML with t(8;21), 37 cases with inv(16)/t(16;16), 83 patients with t(15;17) and 83 AML showing a 11q23/MLL-rearrangement were evaluated. 44 (69.7%), 13 (35.1%), 39 (47%), and 28 (43.1%) cases showed clonal chromosome aberrations in addition, respectively. Therefore, additional chromosomal aberrations are more frequent in all these subgroups than in the AML NPM1mut. Similar to NPM1mut cases +8 (n=2), −X/Y (n=32), +4 (n=2), and del(9q) (n=10) were observed. The only other recurrent additional aberrations was del(11q) (n=2). In inv(16)/t(16;16) we also found +8 (n=5) and −Y (n=1). The only other recurring additional aberrations were +22 (n=6) and del(7q) (n=2). In AML with t(15;17) recurring additional abnormalities were +8 (n=12), −Y (n=3), del(9q) (n=2), ider(17)(q10) t(15;17) (n=7). AML with 11q23/MLL-rearrangement showed +4 (n=2), +8 (n=8), +13 (n=2), +19 (n=4), +21(n=4), +22 (n=2), −Y (n=1). Thus, chromosome aberration in AML NPM1mut share many overlaps to those in AML with recurrent aberrations. Furthermore, the prognostic impact of chromosome aberrations in AML NPM1mut was evaluated. No difference with respect to overall survival (OS) and event-free survival (EFS) was observed between AML NPM1mut with a normal (n=344) and an aberrant karyotype (n=71) (OS at 2 yrs 78% vs. 81%, p=0.969; EFS at 2 yrs 40% vs. 50%, median EFS 544 days vs. 522 days, p=0.253). The FLT3-ITD status was available in 400 cases. 127 (38%) of 334 cases with a normal karyotype showed a FLT3-ITD, while in only 16 (24%) of 66 cases with an aberrant karyotype a FLT3-ITD was observed (p=0.035). While the negative prognostic impact of additional FLT3-ITD was confirmed in our cohort, the presence of chromosome aberrations did not influence prognosis neither in the FLT3-ITD negative nor in the FLT3-ITD positive subgroup. In addition, 31 patients with AML NPM1mut were analyzed by chromosome banding analysis at diagnosis and at relapse (median time diagnosis to relapse: 301 days (range: 71–986). 22 cases (71%) showed a normal karyotype both at diagnosis and relapse. In 4 cases a normal karyotype was observed at diagnosis and an aberrant karyotype at relapse (del(9q) (n=2), t(2;11) (n=1), inv(12) (n=1)). One case with +8 at diagnosis showed +8 also at relapse. One case with +4 at diagnosis showed +4 and additional aberrations at relapse. In 1 case clonal regression was observed (+21 -> normal). One case with an unbalanced 1;3-translocation at diagnosis showed a der(17;18) (q10;q10) at relapse and one case with −Y at diagnosis showed a del(3p) at relapse. In conclusion: 1. Frequency of additional chromosome aberrations is low in AML NPM1mut as compared to other genetically defined WHO entities. 2. The pattern of additional chromosome aberrations is overlapping between the 5 groups analyzed. 3. Chromosome aberrations observed at diagnosis in AML NPM1mut do not influence prognosis in comparison to AML NPM1mut with normal karyotype. 4. The karyotype is stable in most AML NPM1mut patients at diagnosis and at relapse. These results point to chromosomal aberrations occurring in AML NPM1mut as secondary events and further support inclusion of AML NPM1mut as a provisional entity in the new WHO classification.

Genetic and clinical aspects of wood dust related intestinal-type sinonasal adenocarcinoma: a review

Intestinal-type sinonasal adenocarcinoma (ITAC) is a rare epithelial cancer of the nasal cavities and paranasal sinuses. Exposure to wood dust particles is a strong etiological factor making it a professional disease. These tumors are locally aggressive with frequent local recurrences in up to 50% of cases. Metastasis to regional lymph nodes and distant metastasis are less frequent (10%). Invasion of the duramater and local recurrence are frequent and the major cause of death. Standard therapeutic modalities include surgery followed by radiotherapy in advanced stages, sometimes with chemotherapy treatment. The molecular genetic mechanisms underlying the development and progression of this tumor is not understood. Histopathologically, ITAC resembles colorectal adenocarcinoma and have directed early genetic studies to search for similar genetic alterations. Recently, genome-wide studies have identified a recurrent pattern of chromosomal aberrations. This review aims to describe the clinico-pathological characteristics of this relatively unknown tumor and to summarize the knowledge on genetic and chromosomal analyses up to the present time.

Chromosomal Biomarkers for Detection of Human Papillomavirus Associated Genomic Instability in Epithelial Cells of Cervical Cytology Specimens

The goal of this study was to compare how accumulation of chromosomal aberrations in human papillomavirus (HPV)-infected cells correlates with the severity of cervical dysplastic lesions. We assessed the frequency of genomic alterations for 35 different loci in a pilot biopsy study and selected two loci (3q26 and 8q24) with the highest frequency of copy number gains found in high-grade dysplasia and cancer. These probes were labeled with gold and red fluorophores and combined with HPV biotin-labeled probes for subsequent detection using a tyramide signal amplification system with a green fluorophore. Cells that were both HPV positive and chromosomally abnormal were designated as "double-positive cells." Cervical cytology specimens from 235 patients were used for this blinded study. The average number of double-positive cells increased from two cells in patients with a cytological interpretation of atypical squamous cells of undetermined significance to 22 cells in low-grade squamous intraepithelial lesion and 99 cells in high-grade squamous intraepithelial lesion, reflecting an accumulation of chromosomal abnormality with disease progression. Using a cutoff of four or more double-positive cells as the criterion for the presence of a cervical intraepithelial neoplasia 2 or 3 lesion, we demonstrated that low-grade squamous intraepithelial lesion and high-grade squamous intraepithelial lesion cytology specimens with underlying cervical intraepithelial neoplasia 2/3 histology showed positive test results in more than 80% of cases. Correlation of 3q26 and 8q24 aneusomy with concurrent HPV infection may thus serve as a biomarker of genetic instability in HPV-infected cells.