Samuli Hemmer

DNA Copy Number Losses in Human Neoplasms

This review summarizes reports of recurrent DNA sequence copy number losses in human neoplasms detected by comparative genomic hybridization. Recurrent losses that affect each of the chromosome arms in 73 tumor types are tabulated from 169 reports. The tables are available online at http://www.amjpathol.org and http://www. helsinki.fi/ approximately lglvwww/CMG.html. The genes relevant to the lost regions are discussed for each of the chromosomes. The review is supplemented also by a list of known and putative tumor suppressor genes and DNA repair genes (see Table 1, online). Losses are found in all chromosome arms, but they seem to be relatively rare at 1q, 2p, 3q, 5p, 6p, 7p, 7q, 8q, 12p, and 20q. Losses and their minimal common overlapping areas that were present in a great proportion of the 73 tumor entities reported in Table 2 (see online) are (in descending order of frequency): 9p23-p24 (48%), 13q21 (47%), 6q16 (44%), 6q26-q27 (44%), 8p23 (37%), 18q22-q23 (37%), 17p12-p13 (34%), 1p36.1 (34%), 11q23 (33%), 1p22 (32%), 4q32-qter (31%), 14q22-q23 (25%), 10q23 (25%), 10q25-qter (25%),15q21 (23%), 16q22 (23%), 5q21 (23%), 3p12-p14 (22%), 22q12 (22%), Xp21 (21%), Xq21 (21%), and 10p12 (20%). The frequency of losses at chromosomes 7 and 20 was less than 10% in all tumors. The chromosomal regions in which the most frequent losses are found implicate locations of essential tumor suppressor genes and DNA repair genes that may be involved in the pathogenesis of several tumor types.

DNA Copy Number Changes in Thyroid Carcinoma

The genetic changes leading to thyroid cancer are poorly characterized. We studied DNA copy number changes by comparative genomic hybridization (CGH) in 69 primary thyroid carcinomas. In papillary carcinoma, DNA copy number changes were rare (3 of 26, 12%). The changes were all gains, and they were associated with old age (P = 0.01) and the presence of cervical lymph node metastases at presentation (P = 0.08). DNA copy number changes were much more frequent in follicular carcinoma (16 of 20, 80%) than in papillary carcinoma (P < 0.0001), and follicular carcinomas had more often deletions (13/20 versus 0/26, P < 0.0001). Loss of chromosome 22 was common in follicular carcinoma (n = 7, 35%), it was more often seen in widely invasive than in minimally invasive follicular carcinoma (54% versus 0%, P = 0.04), and it was associated with old age at presentation (P = 0.01). In three of the four patients with follicular carcinoma who died of cancer, the tumor had loss of chromosome 22. DNA copy number changes were found in 5 (50%) of the 10 medullary carcinomas studied. Four of these five carcinomas had deletions, and in two of them there was deletion of chromosome 22. Eleven (85%) of the thirteen anaplastic carcinomas investigated had DNA copy number changes, of which five had deletions, and one had deletion of chromosome 22. The most common gains in anaplastic carcinoma were in chromosomes 7p (p22-pter, 31%), 8q (q22-qter, 23%), and 9q (q34-qter, 23%). We conclude that DNA copy number changes are frequent in follicular, medullary, and anaplastic thyroid carcinoma but rare in papillary carcinoma when studied by CGH. Loss of chromosome 22 is particularly common in follicular carcinoma, and it is associated with the widely invasive type.

MET receptor tyrosine kinase sequence alterations in differentiated thyroid carcinoma.

Activating mutations affecting the MET receptor tyrosine kinase are present in several types of human cancer, particularly in papillary renal cell carcinoma. Papillary thyroid carcinomas commonly express high levels of MET mRNA and protein, suggesting that increased MET signaling may be of importance in the molecular pathogenesis of differentiated thyroid carcinoma. To evaluate the role of MET mutations in thyroid carcinoma, we screened MET exons 2 to 21 for mutations in sporadic papillary, follicular, medullary, and anaplastic thyroid carcinomas using denaturing high-performance chromatography. A missense MET sequence alteration T1010I, located in exon 14 encoding for the juxtamembrane domain of MET, was found in 6 (6%) of the 104 thyroid carcinomas examined, whereas all 92 goiter samples had wild-type exon 14 (P = 0.031). Three (6%) of the 53 papillary, 2 (10%) of the 21 follicular, 1 (8%) of the 13 medullary, and none of the 17 anaplastic carcinomas studied had MET(T1010I). Four of the 6 T1010I sequence alterations were present also in the germline. MET protein expression showed no apparent association with the presence of MET(T1010I), and the clinical features of the patients with cancer with MET(T1010I) were similar to those of patients whose cancer did not harbor MET(T1010I). We conclude that MET(T1010I) sequence alteration is relatively frequent in differentiated thyroid carcinoma. The clinical and the molecular pathologic significance of this MET sequence alteration is not known.

Deletion of 11q23 and cyclin D1 overexpression are frequent aberrations in parathyroid adenomas.

Hyperparathyroidism may result from parathyroid hyperplasia or adenoma, or rarely from parathyroid carcinoma. Pericentromeric inversion of chromosome 11 that results in activation of the P:RAD1/cyclin D1 gene and tumor suppressor gene loss have been described as genetic abnormalities in the evolution of parathyroid neoplasms. We studied tissue samples taken from primary parathyroid hyperplasia, parathyroid adenoma, and histologically normal parathyroid tissue by comparative genomic hybridization, fluorescent in situ hybridization, and immunohistochemistry for cyclin D1. DNA copy number changes were infrequent in primary hyperplasia (4 of 24, 17%), but common in adenomas (10 of 16, 63%; P: = 0.0059). The most common change was deletion of the entire chromosome 11 or a part of it, with a minimal common region at 11q23. This change was present in five (31%) adenomas and two (8%) primary hyperplasias. Fluorescent in situ hybridization confirmed the presence of both MEN1 alleles located at 11q13 despite deletion of 11q23 in all three cases studied. Cyclin D1 was overexpressed in six (40%) of the 15 adenomas studied, whereas none of the 27 hyperplasias (P: = 0.0010) nor the five histologically normal tissue samples overexpressed cyclin D1. Either DNA copy number loss or cyclin D1 overexpression was present in 13 (81%) of the 16 adenomas. We conclude that DNA copy number loss and cyclin D1 overexpression are common in parathyroid adenomas. The region 11q23 is frequently lost in parathyroid adenomas and occasionally in parathyroid hyperplasias, and this suggests the possibility that a tumor suppressor gene that is important in their pathogenesis is present on 11q23.

Comparison of benign and malignant follicular thyroid tumours by comparative genomic hybridization.

DNA copy number changes were compared in 29 histologically benign follicular adenomas, of which five were atypical, and 13 follicular carcinomas of the thyroid by comparative genomic hybridization. DNA copy number changes were frequent in adenomas (14 out of 29, 48%). Most changes were gains, and they always involved a gain of the entire chromosome 7 (10 out of 29, 34%); other common gains involved chromosomes 5 (28%), 9 (10%), 12 (24%), 14 (21%), 17 (17%), 18 (14%) and X (17%). Losses were found only in four (14%) adenomas. Two of the five atypical adenomas had DNA copy number losses, and none had gains. Unlike adenomas, gains were rare and losses were frequent in carcinomas. A loss of chromosome 22 or 22q was particularly common in carcinomas (6 out of 13, 46%), whereas a loss of chromosome 22 was found in only two (7%) adenomas, one of which was atypical (P = 0.002). A loss of 1p was also frequent in carcinomas (31%), but gains of chromosomes 5, 7, 12, 14 or X that were common in adenomas were not found. Loss of chromosome 22 or 22q was present in six of the eight widely invasive follicular carcinomas, but in only one of the five minimally invasive carcinomas. We conclude that large DNA copy number changes are common in thyroid adenomas. These changes are strikingly different from those found in follicular carcinomas consisting of few losses and frequent gains, especially those of chromosome 7. A loss of chromosome 22 is common in widely invasive follicular carcinoma.

Differential gene expression in non‐malignant tumour microenvironment is associated with outcome in follicular lymphoma patients treated with rituximab and CHOP

Rituximab in combination with chemotherapy (immunochemotherapy) is one of the most effective treatments available for follicular lymphoma (FL). This study aimed to determine whether differences in gene expression in FL tissue correlate with outcome in response to rituximab and CHOP (cyclophosphamide, doxorubicin, vincristine, prednisone) chemotherapy (R–CHOP). We divided 24 patients into long‐ [time to treatment failure (TTF) >35 months] and short‐term (TTF <23 months) responders, and analysed the gene expression profiles of lymphoma tissue using oligonucleotide microarrays. We used a supervised learning technique to identify genes correlating with outcome, and confirmed the expression of selected genes with quantitative polymerase chain reaction (qPCR) and immunohistochemistry. Among the transcripts with a high correlation between microarray and qPCR analyses, we identified EPHA1, a tyrosine kinase involved in transepithelial migration, SMAD1, a transcription factor and a mediator of bone morphogenetic protein and transforming growth factor‐β signalling, and MARCO, a scavenger receptor on macrophages. According to Kaplan–Meier estimates, high EPHA1, and low SMAD1 and MARCO expression were associated with better progression‐free survival (PFS). Immunohistochemistry showed that EphA1 was primarily localised in granulocytes. In addition, both EphA1 and Smad1 were expressed in vascular endothelia. However, no difference in vasculature was detected between long‐ and short‐term responders. In a validation set of 40 patients, a trend towards a better PFS was observed among patients with high EphA1 expression. We conclude that gene expression in non‐malignant cells contributes to clinical outcome in R–CHOP‐treated FL patients.

Alterations in the suppressor gene PPP2R1B in parathyroid hyperplasias and adenomas

Deletion of chromosome 11q23 is a common alteration in parathyroid adenomas and hyperplasias. A new potential suppressor gene PPP2R1B encoding the beta isoform of the A subunit of the serine/threorine protein phosphatase 2A was recently identified and localized to chromosome 11q23. We performed polymerase chain reaction-based single-strand conformation polymorphism and direct sequencing on six parathyroid hyperplasias and 12 adenomas to evaluate the role of PPP2R1B in the pathogenesis of parathyroid lesions. A previously identified germline G-A transition (GGC-GAC) in codon 90, changing glycine (Gly) to aspartic acid (Asp), was detected in one adenoma. Both the common Gly allele and the variant Asp allele were detected by direct sequencing in the patients somatic cells. We conclude mutations of PPP2R1B are not frequent in parathyroid lesions, and that other genes located at 11q23 may be more closely associated with pathogenesis of parathyroid hyperplasia and adenoma.

DNA copy number losses at 1p32-pter in monozygotic twins concordant for breast cancer

To find similarities that may possibly indicate novel mutations, we performed comparative genomic hybridization (CGH) analysis following degenerate oligonucleotide primed polymerase chain reaction (PCR) for DNA obtained from unique material of breast cancer that developed in monozygotic twin-pairs. Polymerase chain reaction amplification was successful in 12 samples for 11 patients, including 3 pairs. Six samples exhibited DNA copy number changes. Gains (76%) were more frequent than losses (24%). Gains or high-level amplifications in 8q were present in all but 1 of the abnormal cases. Frequent gains were detected with a minimal common overlapping region at 5p (4 cases), at 1q25-qter (3 cases), and at 20q12-qter (2 cases). The most frequent loss, detected in half of the abnormal cases, was at 1p32-pter. One twin-pair showed similar changes in 4 chromosomal locations involving loss of 1p32-pter and gains in 1q25-qter, 5, and 8q.