Pattern Of Chromosomal Alterations Research Articles

PDX1 DNA Methylation Distinguishes Two Subtypes of Pancreatic Neuroendocrine Neoplasms with a Different Prognosis.

DNA methylation is a crucial epigenetic mechanism for gene expression regulation and cell differentiation. Furthermore, it was found to play a major role in multiple pathological processes, including cancer. In pancreatic neuroendocrine neoplasms (PNENs), epigenetic deregulation is also considered to be of significance, as the most frequently mutated genes have an important function in epigenetic regulation. However, the exact changes in DNA methylation between PNENs and the endocrine cells of the pancreas, their likely cell-of-origin, remain largely unknown. Recently, two subtypes of PNENs have been described which were linked to cell-of-origin and have a different prognosis. A difference in the expression of the transcription factor PDX1 was one of the key molecular differences. In this study, we performed an exploratory genome-wide DNA methylation analysis using Infinium Methylation EPIC arrays (Illumina) on 26 PNENs and pancreatic islets of five healthy donors. In addition, the methylation profile of the PDX1 region was used to perform subtyping in a global cohort of 83 PNEN, 2 healthy alpha cell and 3 healthy beta cell samples. In our exploratory analysis, we identified 26,759 differentially methylated CpGs and 79 differentially methylated regions. The gene set enrichment analysis highlighted several interesting pathways targeted by altered DNA methylation, including MAPK, platelet-related and immune system-related pathways. Using the PDX1 methylation in 83 PNEN, 2 healthy alpha cell and 3 healthy beta cell samples, two subtypes were identified, subtypes A and B, which were similar to alpha and beta cells, respectively. These subtypes had different clinicopathological characteristics, a different pattern of chromosomal alterations and a different prognosis, with subtype A having a significantly worse prognosis compared with subtype B (HR 0.22 [95% CI: 0.051–0.95], p = 0.043). Hence, this study demonstrates that several cancer-related pathways are differently methylated between PNENs and normal islet cells. In addition, we validated the use of the PDX1 methylation status for the subtyping of PNENs and its prognostic importance.

Abstract P4-06-04: Effect of genomic heterogeneity on breast cancer progression and metastatic spread

Abstract Purpose: DNA fingerprinting has revealed discordant patterns of chromosomal alterations in primary breast tumors (PBT) compared to matched metastatic axillary lymph node tumors (MLNT); however, whether these genetic differences reflect the timing of dissemination of cells with metastatic potential from the primary tumor, intratumoral heterogeneity of the primary breast tumor, or independent seeding of lymph nodes with metastatic cells remains unclear. Patients and Methods: From the 30 node-positive patients evaluated in this study, allelic imbalance (AI) data was generated using 52 microsatellite markers from 5-19 areas of each PBT as well as from available MLNT. Data were analyzed using pairwise correlations, ANOVA and PHYLIP. Results: The frequency of genomic changes was significantly higher (P<0.001) in PBT areas (13%) than MLNT (9%). No two PBT areas had identical patterns of AI and the percent concordance (PC) of AI events between tumor regions ranged from 0-65% (average 33%). Of the 196 MLNT from 28 patients, PC between MLNT from the same patient ranged from 0-88% (average = 32%). Neither the overall frequency of AI nor the PC differed significantly between sentinel (SLN) and non-SLNs. Phylogenetic analysis revealed that within patients, many MLNT appeared to be descended from different areas of the PBT, but patterns of descent were complex. Conclusions: Both PBT and MLNT are characterized by extensive molecular heterogeneity, MLNT appear to originate from different areas of the PBT, and SLN metastases are not genomically more advanced than non-SLN metastases. These data suggest that metastatic dissemination may be influenced by both spatial and temporal factors, with cells with metastatic potential colonizing lymph nodes throughout the development of the PBT, and that SLN metastases do not appear to be a source of metastatic cells for non-SLN but rather MLNT arise by independent colonization. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P4-06-04.

Genetic changes in nonepithelial ovarian cancer

Nonepithelial ovarian cancers (OCs), including sex cord-stromal tumors (SCSTs) and germ cell tumors (GCTs), are an uncommon subset of OC, together accounting for 10% of all OCs. The etiology of these tumors remains largely unresolved. It is well established that tumorigenesis is the result of multiple genetic alterations driving a normal cell toward a malignant state. Much effort has been made into researching the molecular mechanisms underlying epithelial OC, but far less is known about the genetic changes in SCSTs and GCTs. Recently, a single point missense mutation (C134W) was found in the FOXL2 gene in approximately 95% of adult-type granulosa cell tumors, suggesting a key role for FOXL2 in these tumors. By contrast, the FOXL2 mutation was not found in the juvenile type. DICER1 somatic missense mutations were found in approximately 60% of Sertoli-Leydig tumors. Ovarian GCTs share many morphological features and a similar pattern of chromosomal alterations with testicular GCTs. In the latter, recent genome-wide association studies have identified seven susceptibility loci near KITLG, SPRY4, UKC2, BAK1, DMRT1, TERT and ATF7IP. All of the susceptibility loci detected thus far are all involved in primordial germ cell function or sex determination. TGF-β/BMP and Wnt/β-catenin signaling was absent in dysgerminomas, but present in yolk sac tumors, suggesting intertumoral heterogeneity. In this article, the authors aim to provide an overview of the current knowledge on the possible molecular changes in SCSTs and GCTs of the ovary.

Profiling of GEPNETs

The tumorigenesis of gastrointestinal and pancreatic tumors (GEPNETs) is poorly understood. We need a better understanding of the molecular alterations in GEPNETs to obtain an accurate classification, and it may also provide targets for therapeutic intervention. Purpose of Paper. The purpose of this paper was to critically examine recent advances in the molecular understanding of GEPNETs gained from genome-wide and transcriptome-wide profiling studies. Special emphasis was put on diagnostic, predictive, and therapeutic implications of profiling studies. Results. Pancreatic neuroendocrine tumours (PNETs) were characterised by a distinct pattern of chromosomal alterations and a higher degree of chromosomal instability (CIN) than ileal carcinoids. Subgroups of PNETs and ileal carcinoids were identified on the basis of specific chromosomal alterations. Exome sequencing identified mutations in MEN1, ATRX/DAXX, and mTOR pathway genes as being frequent events in sporadic PNETs. Expression profiles of PNETs and ileal carcinoids were found to be different, and allowed identification of subgroups of tumors, as well discrimination between benign and malignant tumors. The molecular data provided a number of candidate genes and pathways suitable for targeted therapy. For PNETs, candidate targets include BRAF, KRAS, TERT, EGFR, RET, MDM2, IGF, MET/HGF, ANG2, LCK, PDGFRB, AKT-mTOR, and SSTR2. Some of these targets have already been evaluated in clinical trials (mTOR and SSTR2). For ileal carcinoids, significantly fewer candidate targets were provided, including ERBB2 (HER2), RET, APLP1, and Notch. Conclusion. Profiling of GEPNETs is a powerful tool for discovery of novel targets for therapeutic intervention. Further studies, combining genome, epigenome, transcriptome, and proteome data are needed to enable us to identify clinically relevant targets in GEPNETs.

Genomic Alterations Associated with Early Stages of Breast Tumor Metastasis

Molecular studies suggest that acquisition of metastatic potential occurs early in the development of breast cancer; mechanisms by which cells disseminate from the primary carcinomas and successfully colonize foreign tissues are, however, largely unknown. Thus, we examined levels and patterns of chromosomal alterations in primary breast tumors from node-negative (n = 114) and node-positive (n = 115) patients to determine whether specific genomic changes are associated with tumor metastasis. Fifty-two genetic markers representing 26 chromosomal regions commonly altered in breast cancer were examined in laser microdissected tumor samples to assess levels and patterns of allelic imbalance (AI). Real time-PCR (RT-PCR) was performed to determine expression levels of candidate genes. Data was analyzed using exact unconditional and Student's t-tests with significance values of P < 0.05 and P < 0.002 used for the clinicopathological and genomic analyses, respectively. Overall levels of AI in primary breast tumors from node-negative (20.8%) and node-positive (21.9%) patients did not differ significantly (P = 0.291). When data were examined by chromosomal region, only chromosome 8q24 showed significantly higher levels (P < 0.0005) of AI in node-positive primary tumors (23%) versus node-negative samples (6%). c-MYC showed significantly higher levels of gene expression in primary breast tumors from patients with lymph node metastasis. Higher frequencies of AI at chromosome 8q24 in patients with positive lymph nodes suggest that genetic changes in this region are important to the process of metastasis. Because overexpression of c-MYC has been associated with cellular dissemination as well as development of the premetastatic niche, alterations of the 8q24 region, including c-MYC, may be key determinants in the development of lymph node metastasis.

Loss of chromosome 13q is a frequently acquired event in genetic progression of soft tissue sarcomas in the abdominal cavity

Soft tissue sarcomas (STSs) arising in the abdominal cavity constitute a group of aggressive tumours, typically of very large size and with a high recurrence rate in the affected patients. While some distinct genetic etiologies have been described, the genetic background of this tumour group is not well characterised. Here we have assessed gross chromosomal alterations in a series of such tumours obtained from 26 patients. CGH alterations were found in tumours from 23 of the patients (88%), the most frequent being loss of 13q21 (46%) and gain of 17p and/or q (46%). Furthermore, mutations of C-KIT exon 11 were demonstrated in five tumours from four patients, and the two myxoid liposarcomas exhibited a translocation t(12;16)(q13;p11). From the pattern of chromosomal alterations detected, a genetic progression of events was clearly evident in the tumours. Taken together with analysis of subsequent relapses from the same patients, the common CGH alteration +12q13 was suggested to be a relatively early event in the genetic progression, similar to t(12;16)(q13;p11) and C-KIT mutations. Moreover, -1p21-22, -13q21, -14q, -Xp22, +9q34, +17p, +17q, and +20q13 would all represent relative later events. The most consistent alteration was loss of 13q, that was found to target the 13q14-21 and 13q34 regions as determined by CGH and Southern blot analyses. Loss of 13q was identified independently of +12q13 and C-KIT mutation and the patient's sex, and was observed in all common subtypes of STS, suggesting that it is a general and late event in the genetic progression. The findings provide a starting point for further dissection of the target genes involved in development of STSs in the abdominal cavity.

Robust classification of renal cell carcinoma based on gene expression data and predicted cytogenetic profiles.

Renal cell carcinoma (RCC) is a heterogeneous disease that includes several histologically distinct subtypes. The most common RCC subtypes are clear cell, papillary, and chromophobe, and recent gene expression profiling studies suggest that classification of RCC based on transcriptional signatures could be beneficial. Traditionally, however, patterns of chromosomal alterations have been used to assist in the molecular classification of RCC. The purpose of this study was to determine whether it was possible to develop a classification model for the three major RCC subtypes that utilizes gene expression profiles as the bases for both molecular genetic and cytogenetic classification. Gene expression profiles were first used to build an expression-based RCC classifier. The RCC gene expression profiles were then examined for the presence of regional gene expression biases. Regional expression biases are genetic intervals that contain a disproportionate number of genes that are coordinately up- or down-regulated. The presence of a regional gene expression bias often indicates the presence of a chromosomal abnormality. In this study, we demonstrate an expression-based classifier can distinguish between the three most common RCC subtypes in 99% of cases (n = 73). We also demonstrate that detection of regional expression biases accurately identifies cytogenetic features common to RCC. Additionally, the in silico-derived cytogenetic profiles could be used to classify 81% of cases. Taken together, these data demonstrate that it is possible to construct a robust classification model for RCC using both transcriptional and cytogenetic features derived from a gene expression profile.

Clinical significance of allelotype profiling for urothelial carcinoma

Objectives To perform a global loss of heterozygosity (LOH) analysis on a cohort of urothelial carcinoma to investigate the clinical implication of specific chromosomal loss. Allelic deletions detected as LOH have been used to study the markers for carcinogenesis. Methods We examined the allelic loss on 14 chromosomal regions in a total of 71 cases of urothelial carcinoma. The results were analyzed in relation to biologic indicators of urothelial carcinoma and the clinical outcome with a mean follow-up of 101 months. Results The incidence of LOH in order of frequency was 9p (54.9%), 9q (49.3%), 13q (40.8%), 14q (40.8%), 10q (39.4%), 17p (39.4%), 8p (38.0%), 21q (36.6%), 11p (31.0%), 18q (23.9%), 4q (21.1%), 3p (16.9%), 6q (14.1%), and 1q (8.5%). Positive association with one of the indicators was observed in 3p, 9p, 9q, 10q, 14q, and 18q. The chromosomes that correlated with two biologic indicators were 4q, 6q, 11p, 17p, and 21q. Univariate analysis found that patients having combined 9p and 14q deleted tumors had particularly poor long-term survival compared with those with other patterns of chromosomal alterations ( P = 0.01). In the multivariate model, nonpapillary tumors had a greater risk of recurrence, and stage classification was the only important indicator in predicting patient survival ( P = 0.04). Conclusions LOH assessment does not provide independent prognostic value compared with stage classification. However, chromosomes 4q, 6q, 9p, 11p, 14q, 17p, and 21q may harbor important tumor suppressor genes involved in the progression of urothelial carcinogenesis.

DNA amplification on chromosome 7q in squamous cell carcinoma of the tongue.

Squamous cell carcinoma of the head and neck exhibit a highly variable picture of chromosomal aberrations. In the present study the clearly defined anatomical region of the tongue was analyzed for potentially specific patterns of chromosomal alterations. Fresh tumor samples from 18 patients afflicted by squamous cell carcinoma of the tongue constituted the clinical basis of the present investigation. The tumor samples were analyzed on the basis of comparative genomic hybridization (CGH), a molecular cytogenetic FISH-approach. Gains in DNA copy numbers were detected as the predominant imbalance on chromosomes 7q (9/18), 3q (48/18), 16p (7/18) and 20q (7/18). The regions of minimal overlap on these chromosomes were mapped to 7q11.2q11.3 and 3q26. A conspicuous finding was the frequent detection of amplifications in the 7q11 region. Gains in the 7q region have been rarely reported in CGH studies of tumors derived from different regions of the head and neck. Amplifications on 7q could thus be specifically linked with the tongue region and could correlate with specific clinical factors of this tumor entity.

Chromosomal Alterations in Paired Gastric Adenomas and Carcinomas

Gastric adenoma is a precancerous lesion of the stomach and its malignant transformation is thought to result from accumulative series of gene alterations. The aim of this study was to determine the pattern of chromosomal changes during gastric carcinogenesis. Pairs of adenoma and carcinoma tissues from 15 gastrectomy cases containing both adenomas and carcinomas in the same (adjacent pairs, 6 cases) and different (non-adjacent pairs, 9 cases) lesions, were analyzed for chromosomal alterations of 39 non-acrocentric chromosomal arms by comparative genomic hybridization (CGH). CGH analysis identified frequent chromosomal alterations in most of the gastric adenomas (14/15, 93%) and all of the carcinomas. The mean number of chromosomal alterations was higher in carcinoma (5.5 for adenoma and 11.7 for carcinoma; P = 0.006, by nonparametric Wilcoxon's test). Losses on the short arm of chromosome 17 were most common in both adenomas (43%) and carcinomas (67%). The pattern of chromosomal alterations in paired gastric adenomas and carcinomas showed greater similarity compared to the non-case pairs and this similarity was increased in the adjacent pairs. Deletion mapping analysis on chromosome 17p also demonstrated that the conserved deletion area was more frequent in the adjacent pairs. Among these 6 adjacent pairs, all had common deletion areas. In contrast, among the 9 non-adjacent pairs, 2 (22%) had common area of deletion, 5 (56%) showed deletion only in the carcinoma, and the remaining 2 (22%) had no deletion on 17p, suggesting diverse genetic changes might be involved in the multiple tumor formation. Our results that common clonal genetic changes between adjacent pairs of gastric adenomas and carcinomas and accumulated genetic changes in the carcinomas provide evidences for the stepwise mode of gastric carcinogenesis through the accumulation of a series of genetic alterations.

Human papilloma virus status and chromosomal imbalances in primary cervical carcinomas and tumour cell lines

Human papilloma virus (HPV) infection is the crucial step in the initiation of cervical carcinomas. In addition, HPV18 has been implicated in tumour progression and adverse clinical outcome. We determined the HPV types in 12 primary cervical carcinomas and 12 cell lines and compared the findings with the comparative genetic hybridisation (CGH) pattern of chromosomal alterations. The most frequent alteration was the deletion at 3p14 followed by the loss of 2q34-q36 along with 3q gain. High risk HPV types were detected in all samples except one primary tumour. In contrast to the normal distribution, HPV18 was present in 75% of cases including all cell lines. The cell lines carried a higher number of genetic alterations and a different CGH pattern for several chromosomes than the primary tumours, despite microdissection. Purely HPV18 positive cases indicated a high incidence of imbalances at specific loci with peaks of the histogram coinciding with known HPV integration sites. The study suggests that HPV infection is associated with a recurrent pattern of chromosomal changes in cervical carcinomas and that the development and progression of these alterations is triggered by integration into the host genome.

Deletion of chromosome 10q--a marker for metastasis of head-neck carcinomas?

Comparative genomic hybridization (CGH) was applied to squamous cell carcinomas of the head and neck to define genetic alterations that are associated with the metastatic phenotype. CGH is a molecular cytogenetic method allowing the comprehensive analysis of a tumor genome for chromosomal imbalances. In total, 23 primary squamous cell carcinomas without evidence of metastasis formation and 20 lymph node metastases were investigated. Prevalent changes observed in more than 50% of the primary tumors included deletions on chromosomes 3p, 4p/q, 5q, 6q, 9p, 11q, 13q, and 18q, and DNA overrepresentations on chromosomes 1p, 3q, 5p, 8q, 9q, 11q13, 16p, 17q, 19p, 20q, and 22q. To evaluate the differences between both groups we used a histogram representation, calculation of a difference histogram, and statistical analysis. The analysis revealed that the lymph node metastases were frequently characterized by deletions on chromosomes 10, 11, and 14. In particular, DNA loss of the chromosomal bands 5p12, 10p11.2-12, 10q21, 10q22-23, 10q24-26, 11p13-14, 11q24-25, and 14q22-24 were significantly associated with metastases formation. The statistical analysis indicated that particularly the deletions on chromosome 10q were highly significant markers for the incidence of lymph node metastases. Our data indicate that tumor phenotypes are determined by patterns of chromosomal alterations, and that 10q deletions may predict the metastatic phenotype in head and neck squamous cell carcinomas.

An Integrated BAC/PAC Resource for identifying chromosomal abnormalities in solid tumors

Cancer is a single disease and it is a hundred diseases. To understand and treat cancer, we must know the foe: its morphology, its deregulated genes and pathways, and the patterns of chromosomal alterations that are associated with malignant transformation. In order to do this, we need molecular tools to link the visible alterations of clinical disease with the underlying genes and emerging genome sequences. Our laboratory has developed such ‘tools’ by creating an integrated BAC/PAC Resource for the entire genome by using high resolution fluorescence in situ hybridization (FISH) followed by integration with the genetic, STS and radiation hybrid maps. This resource now covers 30% of the entire human genome, and contains >1,000 STS-linked BACs (Korenberg et al., 1999) and >6,000 mapped BAC/PAC clones.The goal is to construct an integrated overlapping array of molecular cytogenetic markers that covers 90% of the genome for rapid detection of cancer breakpoints and subtle aneuploidies using a combination of FISH and array technologies. Using this resource, we have already identified an oncogene associated with thyroid tumors (Chen et al, 1998). The ultimate intent will be to define characteristic diagnostic signatures for staged tumors as well as prognostic signatures to sensitively reflect the response to treatment. We propose that the generation of such a resource would best be accomplished by integration of current efforts. This will speed the understanding of cancers and ultimately the treatment of our aging population.

Detection of chromosomal aberrations in seminomatous germ cell tumours using comparative genomic hybridization

Comparative genomic hybridization (CGH) was used to evaluate tissue specimens from 16 seminomas in order to elucidate the pathogenesis of germ cell tumours in males. A characteristic pattern of losses and gains within the entire genomes was detected in 94% of the seminomas by comparing the ratio profiles of the tumours with a standard of cytogenetically normal genomic DNA. Losses represented 43% of the total number of alterations often affecting chromosomes and chromosome arms 4, 5, 11, 13q, and 18q. Gains amounted to 57% and were often observed on 1q, 7, 8, 12, 14q, 15q, 21q, and 22q. Aberrations of 12p and 21q appeared most consistently. Results from CGH analysis displayed no relationship to the clinical stages of the malignancy. Some rare aberrations appeared, however, only in clinical stage II and in tumours showing relapse in the contralateral testis following orchiectomy, although the alterations were not present in all of the tumours in question. Losses of 16q13-21 and gains of 9q22.1-22.2 were demonstrated in both groups, while loss of 16p12 and gains of 6p21 and 6q23.3-24 were detected in the latter group as well. In conclusion, a specific pattern of chromosomal alterations was demonstrated in the seminomas by improved detection criteria, which increased specificity and sensitivity. The rare aberrations, which appeared only in tumours in improved detection criteria, which increased specificity and sensitivity. The rare aberrations, which appeared only in tumours in clinical stage II and relapsed tumours, may be linked to tumour progression, invasiveness, and bilateral disease.