Maija Tarkkanen

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 sequence copy number increase at 8q: A potential new prognostic marker in high-grade osteosarcoma

Histologic response to chemotherapy is currently the best prognostic parameter in high‐grade osteosarcoma but it can be evaluated only after several weeks of chemotherapy. Thus a prognostic parameter known at the time of diagnosis would be of great clinical benefit. In the present study, we present the results of 31 primary high‐grade osteosarcomas analyzed by comparative genomic hybridization (CGH). CGH allows for genome‐wide screening of a tumor by detecting alterations in DNA sequence copy number. The most frequent aberrations were copy number increases at 1q21 in 58% of the tumors and at 8q (8q21.3‐q22 in 52% and 8cen‐q13 in 45%), followed by copy number increases at 14q24‐qter (35%) and Xp11.2‐p21 (35%). The most common losses were detected at 6q16 (32%) and 6q21‐q22 (32%). Patients with a copy number increase at 8q21.3‐q22 and/or at 8cen‐q13 had a statistically significant poor distant disease‐free survival (p = 0.003) and showed a trend toward short overall survival (p = 0.04). Patients with a copy number increase at 1q21 showed a trend toward short overall survival (p = 0.04). Thus, specific genetic aberrations detected at the time of the diagnosis could be used in prognostic evaluation of high‐grade osteosarcoma. Int. J. Cancer (Pred. Oncol.) 84:114–121, 1999.

LKB1 Somatic Mutations in Sporadic Tumors

Germline mutations of LKB1/Peutz-Jeghers syndrome gene predispose carriers to hamartomatous polyposis of the gastrointestinal tract as well as to cancer of different organ systems. Although Peutz-Jeghers syndrome patients frequently present with neoplasms of the colon, stomach, small intestine, pancreas, breast, ovaries, and cervix, somatic mutations appear to be rare in the sporadic tumor types thus far studied (colorectal, gastric, testicular, and breast cancers). To evaluate whether somatic mutations of LKB1 contribute to the tumorigenesis of yet unstudied tumor types, we screened 14 cell lines and 129 tumor specimens from different cancers for a genetic defect in LKB1. Six melanoma and eight myeloma cell lines were scrutinized for LKB1 somatic mutations by genomic sequencing. No changes were found in the coding LKB1 sequence and exon/intron boundaries. Next, we analyzed 12 pancreatic, 8 gastric, 12 ovarian granulosa cell, 26 cervical, 28 lung, 24 soft tissue, and 19 renal tumors by single-strand conformational polymorphism analysis. Three changes in LKB1 coding nucleotide sequence were identified. One base pair deletion at A957 and G958 substitution by T occurred in a cervical adenocarcinoma sample, resulting in a frameshift and premature stop codon at position 335. Substitution of A581 by T occurred in a lung adenocarcinoma sample, resulting in the change of aspartic acid at position 194 to valine. A loss of another allele was detected in this sample. One silent change, C1257T, was found in a pancreatic carcinoma sample. The changes were not present in the matched normal tissue DNA samples. Our results suggest that mutational inactivation of LKB1 is a rare event in most sporadic tumor types.

Gene amplifications in osteosarcoma—CGH microarray analysis

Little is known about the genomic alterations underlying osteosarcoma. We performed a genomewide high‐resolution gene copy number analysis of 22 osteosarcoma samples using comparative genomic hybridization on a cDNA microarray that contained cDNA clones of about 13,000 genes. Nineteen of the 22 cases had amplifications that on average spanned more than 1 Mb and contained more than 10 genes. Numerous regions of gain and loss were identified, and their boundaries were defined at high resolution. Novel amplicons were found at 14q11, 17q25, and 22q11–q13. Earlier‐known large amplified regions were detected at 12q11–q15, 8q24, 6p12–p13, and 17p11–p13 in 8, 6, 5, and 4 of the 22 samples, respectively. Amplification of 12q was observed more frequently (36% of the cases) than previously reported. Previously known small amplicons at 1p34–p36, 1q21, 19q13, and 21q22 were seen in at least three cases. Our results implicate TOM1L2 and CYP27B1 as having roles as novel targets for the 17p and 12q amplicons, respectively. Details (www.helsinki.fi/cmg) of the amplified genes in each amplicon provide valuable raw data for further in silico studies.

Novel findings in gene expression detected in human osteosarcoma by cDNA microarray.

cDNA microarray analysis was used to screen for gene expression alterations in human osteosarcoma cell lines. The analysis using three cell lines revealed changes in the expression of several genes in comparison with normal human osteoblasts. Among the 5,184 sequences that were analyzed, 35 showed aberrant expression in all the cell lines. Eight of these showed overexpression and 27 underexpression compared to their expression levels in osteoblasts. The most highly up-regulated genes included heat shock protein 90beta and polyadenylate-binding protein-like 1. Commonly down-regulated genes included fibronectin 1 and thrombospondin 1. RT-PCR was used to verify these changes in the cell lines and in three primary osteosarcoma samples. This study shows that (1) gene expression pattern in osteosarcoma cell lines differs considerably from normal osteoblasts, (2) osteosarcoma cell lines can be used as a model system to detect novel gene expression alterations present in primary tumors, (3) the overexpression of heat shock protein 90beta and polyadenylate-binding protein-like 1, and (4) the down-regulation of fibronectin 1 and thrombospondin 1 may play a role in the development and/or progression of osteosarcoma. This study indicates that microarray-based expression surveys may be used to establish the molecular fingerprint of osteosarcoma, however, larger cDNA chips and more tumor specimens are required to define the clinically relevant gene expression patterns.

Amplification of 17p11.2∼p12, including PMP22, TOP3A, and MAPK7, in high-grade osteosarcoma

Amplification of region 17p11.2 approximately p12 has been found in 13%-29% of high-grade osteosarcomas, suggesting the presence of an oncogene or oncogenes that may contribute to their development. To determine the location of these putative oncogenes, we established 17p11.2 approximately p12 amplification profiles by semiquantitative PCR, using 15 microsatellite markers and seven candidate genes in 19 high-grade osteosarcomas. Most of the tumors displayed complex amplification profiles, with frequent involvement of marker D17S2041 in 17p12 and TOP3A in 17p11.2 and, in some cases, very high-level amplification of PMP22 and MAPK7 in 17p11.2. Our findings suggest that multiple amplification targets, including PMP22, TOP3A, and MAPK7 or genes close to these candidate oncogenes, may be present in 17p11.2 approximately p12 and thus contribute to osteosarcoma tumorigenesis.

Impact of the smallest surgical margin on local control in soft tissue sarcoma

The aim was to review a single‐institution experience of a prospective treatment protocol for soft tissue sarcoma of the extremity and trunk wall, with particular focus on the smallest surgical margin leading to local control.

Comparison of genetic changes in primary sarcomas and their pulmonary metastases.

The aims of the present study were to compare genetic aberrations in primary sarcomas and their pulmonary metastases and to explore the pathways associated with disease spreading. The primary tumor and its subsequent pulmonary metastasis of 22 patients were analyzed by comparative genomic hybridization. All samples were obtained before the initiation of chemo‐ or radiotherapy. The mean total number of aberrations per tumor was 7.6 (range, 0–17) in primary tumors and 7.5 (range, 0–19) in metastases. The mean numbers of high‐level amplifications per tumor were similar (0.32 in primary tumors and 0.36 in metastases). The frequencies of the most common aberrations were relatively similar in primary tumors and metastases: the most frequent gain affected 1q (minimal common regions 1q21–q23 in 36% of primary tumors and 1q21 in 45% of metastases). The most frequent losses were detected at 9p (9p22–pter in 32% of primary tumors and 9p21–pter in 32% of metastases), 10p (10p11.2–p12 in 41% of primary tumors and 10p11.2–pter in 32% of metastases), 11q (11q23–qter in 36% of primary tumors and 32% of metastases), and 13q (13q14–q21 in 45% of primary tumors and 50% of metastases). No aberrations specific to metastases were detected. An increase in the total number of changes during progression was a predominant feature in a majority of these paired samples. Also, the number of differences in the genetic profile outnumbered common changes in a majority of the samples. However, despite the heterogeneous and numerous changes, all pairs with aberrations in both specimens had some shared alterations in both samples. Genes Chromosomes Cancer 25:323–331, 1999.

Incidence, epidemiology and treatment results of osteosarcoma in Finland - a nationwide population-based study

Abstract Background. Patients diagnosed with osteosarcoma in Finland during 1991–2005 were retrospectively analyzed in a nationwide, population-based study. We focused on the incidence, treatment and outcome of osteosarcoma patients. We also evaluated the value of known prognostic parameters. Material and methods. Osteosarcomas were retrieved from the files of the national Finnish Cancer Registry. Only patients with histologically confirmed osteosarcoma were included in the analysis. Histological review was performed. Results. The study consists of 144 osteosarcoma patients with a mean follow-up of 9.8 years for survivors. Mean annual incidence of histologically confirmed osteosarcoma was 1.8 new osteosarcomas per million. The 10-year sarcoma-specific survival for the whole population was 63% and 73% for patients with local disease at presentation. Overall limb-salvage rate was 73% and local control was 84% for patients with a peripheral tumor. Development of local recurrence and major deviation from the chemotherapy protocol were significant adverse factors for sarcoma-specific survival in multivariate analysis. Conclusion. The present nationwide and population-based study is our second report of treatment and prognosis of osteosarcoma in Finland. With modern chemotherapy the prognosis of local osteosarcoma has improved in Finland from 47% during 1971–1980 and 65% during 1981–1990 at five years to the present 73% during 1991–2005 at 10 years. The 10-year sarcoma-specific survival of 73% is excellent and comparable to results reported with contemporary treatment protocols in high-volume centers. However, improvement in limb-salvage rate and local control probably requires centralization of treatment of this rare disease.

Soft tissue sarcoma - a population-based, nationwide study with special emphasis on local control

Abstract Background. A prospective diagnostics and treatment protocol for extremity and trunk wall soft tissue sarcoma (STS) was introduced by the Scandinavian Sarcoma Group in 1986 and it was also widely adopted in Finland. We have updated the protocol and made it more detailed at the Helsinki University Central Hospital. We retrospectively compared diagnostics and treatment of STS in a nationwide population-based material to this protocol with special emphasis on local control. Methods. Data for 219 patients with an STS of extremity or trunk wall diagnosed during 1998–2001 was retrieved from the nationwide Finnish Cancer Registry. Histologic review was performed. Treatment centres were divided into high-, intermediate- and low-volume centres based on the number of patients with final surgery during the study period. Results. Significantly more patients were operated with a preoperative histological or cytological diagnosis at high-volume centres. No preoperative diagnosis was a strong predictor for the patient to undergo more than one operation (p < 0.0001). Wide surgical margin was achieved more often at high-volume centres, but in all centre categories a considerable percentage of patients with inadequate surgical margin did not receive adjuvant radiation therapy. Local control at five years was 82% at high-volume centres, 61% at intermediate-volume centres treating highest percentage of deep tumours and 69% at low-volume centres (p = 0.046). Local control improved as the number of patients operated (surgical volume of the centre) increased. Conclusion. The present quality-control study is the first nationwide population-based study to assess diagnostics and treatment of STS. When referred to a specialised sarcoma centre even patients with inadequate surgery can achieve good local control. STS is a rare cancer and its treatment should be centralised in Finland, which has 5.4 million inhabitants and approximately 100 new STSs of extremities and trunk wall annually.