Identification of chromosome aberrations in sporadic microsatellite stable and unstable colorectal cancers using array comparative genomic hybridization.

Colorectal cancer (CRC) is one of the most common cancers in Denmark and in the western world in general, and the prognosis is generally poor. According to the traditional molecular classification of sporadic colorectal cancer, microsatellite stable (MSS)/chromosome unstable (CIN) colorectal cancers constitute approximately 85% of sporadic cases, whereas microsatellite unstable (MSI) cases constitute the remaining 15%. In this study, we used array comparative genomic hybridization (aCGH) to identify genomic hotspot regions that harbor recurrent copy number changes. The study material comprised fresh samples from 40 MSS tumors and 20 MSI tumors obtained from 60 Danish CRC patients. We identified five small genomic regions (<15 megabases) exhibiting recurrent copy number loss, which, to our knowledge, have not been reported in previously published aCGH studies of CRC: 3p25.3, 3p21.2-p21.31, 5q13.2, 12q24.23-q24.31, and 12q24.23-q24.31. These regions contain several potentially important tumor suppressor genes that may play a role in a significant proportion of both sporadic MSS CRC and MSI CRC. Furthermore, the generated aCGH data are in support of the recently proposed classification of sporadic CRC into MSS CIN+, MSI CIN-, MSI CIN+, and MSS CIN- cancers.

Resumption of mitosis in frozen-thawed embryos is not related to the chromosomal constitution.

OBJECTIVE: To study the relation between the resumption of mitosis after thaw and chromosomal constitution in frozen-thawed embryos. In addition, to evaluate the correlation among the three parameters of resumption of mitosis after thaw, postthaw blastomere loss, and multinucleation. DESIGN: Frozen-thawed embryos were morphologically evaluated at thaw and after 24 hours of culture. Then, fluorescence in situ hybridization (FISH) analysis, including enumeration of 13 chromosomes, was performed by using a combination of peptide nucleic acid and DNA probes. SETTING: In vitro fertilization laboratory. PATIENT(S): Forty IVF and/or intracytoplasmic sperm injection patients. INTERVENTION(S): Embryo thawing, morphological evaluation, and fluorescence in situ hybridization analysis for aneuploidy screening. MAIN OUTCOME MEASURE(S): Resumption of mitosis, blastomere loss, multinucleation, and chromosome enumeration. RESULT(S): No difference was observed in the chromosomal constitution of embryos with and without resumption of mitosis. Neither was the postthaw blastomere loss connected to the chromosomal constitution. The resumption of mitosis was not associated with postthaw loss of blastomeres or with multinuclearity. CONCLUSION(S): Resumption of mitosis and blastomere loss of frozen-thawed embryos is not related to chromosome aberrations in the embryo. Further, the resumption of mitosis is not correlated with multinucleation. However, the high incidence of multinucleated embryos after thawing indicates that the freezing and thawing procedure may affect this condition.

Non-disjunction of chromosome 13.

We performed a molecular study with 21 microsatellites on a sample of 82 trisomy 13 conceptuses, the largest number of cases studied to date. The parental origin was determined in every case and in 89% the extra chromosome 13 was of maternal origin with an almost equal number of maternal MI and MII errors. The latter finding is unique among human autosomal trisomies, where maternal MI (trisomies 15, 16, 21, 22) or MII (trisomy 18) errors dominate. Of the nine paternally derived cases five were of MII origin but none arose from MI errors. There was some evidence for elevated maternal age in cases with maternal meiotic origin for liveborn infants. Maternal and paternal ages were elevated in cases with paternal meiotic origin. This is in contrast to results from a similar study of non-disjunction of trisomy 21 where paternal but not maternal age was elevated. We find clear evidence for reduced recombination in both maternal MI and MII errors and the former is associated with a significant number of tetrads (33%) that are nullichiasmate, which do not appear to be a feature of normal chromosome 13 meiosis. This study supports the evidence for subtle chromosome-specific influences on the mechanisms that determine non-disjunction of human chromosomes, consistent with the diversity of findings for other trisomies.

Laser microdissection and microsatellite analysis of colorectal adenocarcinomas.

BACKGROUND: Microsatellite instability (MSI) is an important marker in colorectal cancer. The analysis may be difficult if the tumour is heterogeneous or only scarce material is available. The aim of this study was to apply laser microdissection (LMD) to MSI analysis in an attempt to allow diagnosis in these situations. MATERIALS AND METHODS: Twenty-two primary tumours and eight lymph node metastases from twenty patients were laser microdissected and MSI analysis was performed with an optimised multiplex PCR. Differences in allelic size between tumour and blood were evaluated to determine the MSI status. RESULTS: The method proved efficient in as little as 4,000 microm3 formalin-treated and paraffin-embedded tumour tissue. The result of microsatellite analysis was independent of sample location in the primary tumour and its metastasis. CONCLUSION: LMD followed by a multiplex PCR is a useful method for MSI analysis in cases of tumour heterogeneity and scarce tumour material.

Systematic re-examination of carriers of balanced reciprocal translocations: a strategy to search for candidate regions for common and complex diseases.

Balanced reciprocal translocations associated with genetic disorders have facilitated the identification of a variety of genes for early-onset monogenic disorders, but only rarely the genes associated with common and complex disorders. To assess the potential of chromosomal breakpoints associated with common/ complex disorders, we investigated the full spectrum of diseases in 731 carriers of balanced reciprocal translocations without known early-onset disorders in a nation-wide questionnaire-based re-examination. In 42 families, one of the breakpoints at the cytogenetic level concurred with known linkage data and/or the translocation co-segregated with the reported phenotype, for example, we found a significant linkage (lod score=2.1) of dyslexia and a co-segregating translocation with a breakpoint in a previously confirmed locus for dyslexia. Furthermore, we identified 441 instances of at least two unrelated carriers with concordant breakpoints and traits. If applied to other populations, re-examination of translocation carriers may identify additional genotype-phenotype associations, some of which may be novel and others that may coincide with and provide additional support of data presented here.

An excess of chromosome 1 breakpoints in male infertility.

In a search for potential infertility loci, which might be revealed by clustering of chromosomal breakpoints, we compiled 464 infertile males with a balanced rearrangement from Mendelian Cytogenetics Network database (MCNdb) and compared their karyotypes with those of a Danish nation-wide cohort. We excluded Robertsonian translocations, rearrangements involving sex chromosomes and common variants. We identified 10 autosomal bands, five of which were on chromosome 1, with a large excess of breakpoints in the infertility group. Some of these could potentially harbour a male-specific infertility locus. However, a general excess of breakpoints almost everywhere on chromosome 1 was observed among the infertile males: 26.5 versus 14.5% in the cohort. This excess was observed both for translocation and inversion carriers, especially pericentric inversions, both for published and unpublished cases, and was significantly associated with azoospermia. The largest number of breakpoints was reported in 1q21; FISH mapping of four of these breakpoints revealed that they did not involve the same region at the molecular level. We suggest that chromosome 1 harbours a critical domain whose integrity is essential for male fertility.