Faculty Bibliography

Activation of oncogenic Kras in murine lung leads to the development of numerous small adenomas, only some of which progress over time to overt adenocarcinoma. Thus, although Kras is the initiating oncogene, it is likely that secondary genetic events are required for progression from adenoma to adenocarcinoma. Some of these secondary events may also be important in human lung adenocarcinoma. By comparing gene expression profiles with DNA copy number changes, we sought to identify genes that play key roles in tumor progression in this model. Gene expression profiling revealed significant heterogeneity among the tumor samples. In 27% of the tumors analyzed, whole- or sub-chromosome duplications or deletions in one or more chromosomes were seen. Recurrent duplications were seen on chromosomes 6, 8, 16, and 19, whereas chromosomes 4, 11, and 17 were frequently lost. Notably, focal amplifications or deletions were not seen. Despite the lack of focal amplification, we showed that chromosome duplication has a measurable effect on gene expression that is not uniform across the genome. We identified a group of genes whose gene expression was highly correlated with changes in DNA copy number. These highly correlated genes were enriched for gene ontology categories involved in the DNA damage response and telomere maintenance.

The short report will be focused on the genetic basis and possible mechanisms of tumorigenesis, common types of cancer, the importance of genetic diagnosis of cancer, and the methodology of cancer genetic diagnosis. They will also review presymptomatic testing of hereditary cancers, and the application of expression profiling to identify patients likely to benefit from particular therapeutic approaches.

Trisomy of the long arm of chromosome 20 is rare. We describe an 18-month-old male who was born at 36 weeks via Caesarian section after an uneventful pregnancy. During the newborn period he was found to have a right-sided cleft lip and cleft palate, hypertelorism, strabismus and mildly over-folded ears with cupping. Cardiovascular examination was consistent with the diagnosis of severe aortic coarctation, which was confirmed by echocardiogram. Additionally, hypothyroidism was diagnosed. Neurological evaluation at 18 months revealed a hypotonic infant with delayed acquisition of motor milestones. Cytogenetic analysis showed additional material on the long arm of chromosome 20, confirmed by fluorescence in situ hybridization (FISH) analysis as being of chromosome 20 origin. Because of the indistinct GTG-banding pattern it was not possible to distinguish between a proximal [dup(20)(q11.2q13.1)] or distal duplication [dup(20)(q13.1q13.3)]. To further define the duplication we used array comparative genomic hybridization (CGH) which demonstrated a 7.8 Mb interstitial duplication in distal 20q. Thus, the proband's karyotype was interpreted as 46,XY,dup(20)(q13.2q13.2). The proband is the first reported case of a pure duplication of this region. This case further highlights the utility of array CGH in characterizing aneusomies and, in particular, for accurate breakpoint designation and quantitation of ambiguous rearrangements.

Absence of the corpus callosum is often associated with cognitive deficits, autism, and epilepsy. Using a genomic microarray, the authors analyzed DNA from 25 patients with radiographically confirmed callosal anomalies and identified three patients with de novo copy number changes in chromosome regions 2q37, 6qter, and 8p. Chromosomal deletions and duplications may be a relatively common cause of cerebral malformations.

Interstitial deletions of chromosome 12q are rare, with only 11 reported cases in the literature. We recently described two cases with cytogenetically identical interstitial deletions of the long arm of chromosome 12. Here, we report on a third patient, a 26-month-old male with a cytogenetically-identical interstitial deletion: 46,XY,del(12)(q21.2q22). Phenotypic features of this male proband included craniofacial and ectodermal anomalies, genitourinary anomalies, minor cardiac abnormalities, mild ventriculomegaly on brain MRI, hyperopia, and developmental delay. To further define the extent of the chromosomal aberration, microarray-based comparative genomic hybridization (array CGH) analysis was performed and the array data was compared to one of our previously reported cases. Although cytogenetic analysis of the two patients was concordant, molecular analysis by array CGH revealed that the patients had discordant distal breakpoints. The determination of molecular breakpoints and phenotypic analyses in these two patients, in conjunction with previously reported cases, leads us to propose a 12q deletion phenotype and a possible genetic locus for hyperkeratosis pilaris/ulerythema ophryogenes.

Therapy-induced cancers are a severe complication of genotoxic therapies. We used heterozygous Nf1 mutant mice as a sensitized genetic background to investigate tumor induction by radiation (RAD) and cyclophosphamide (CY). Mutagen-exposed Nf1(+/-) mice developed secondary cancers that are common in humans, including myeloid malignancies, sarcomas, and breast cancers. RAD cooperated strongly with heterozygous Nf1 inactivation in tumorigenesis. Most of the solid tumors showed loss of the wild-type Nf1 allele but retained two Trp53 alleles. Comparative genomic hybridization demonstrated distinct patterns of copy number aberrations in sarcomas and breast cancers from Nf1 mutant mice, and tumor cell lines showed deregulated Ras signaling. Nf1(+/-) mice provide a tractable model for investigating the pathogenesis of common mutagen-induced cancers and for testing preventive strategies.

BACKGROUND: Fryns syndrome (FS) is the commonest autosomal recessive syndrome in which congenital diaphragmatic hernia (CDH) is a cardinal feature. It has been estimated that 10% of patients with CDH have FS. The autosomal recessive inheritance in FS contrasts with the sporadic inheritance for the majority of patients with CDH and renders the correct diagnosis critical for accurate genetic counselling. The cause of FS is unknown. METHODS: We have used array comparative genomic hybridisation (array CGH) to screen patients who have CDH and additional phenotypic anomalies consistent with FS for cryptic chromosome aberrations. RESULTS: We present three probands who were previously diagnosed with FS who had submicroscopic chromosome deletions detected by array CGH after normal karyotyping with G-banded chromosome analysis. Two female infants were found to have microdeletions involving chromosome band 15q26.2 and one male had a deletion of chromosome band 8p23.1. CONCLUSIONS: We conclude that phenotypes similar to FS can be caused by submicroscopic chromosome deletions and that high resolution karyotyping, including array CGH if possible, should be performed prior to the diagnosis of FS to provide an accurate recurrence risk in patients with CDH and physical anomalies consistent with FS.

Bacterial Artificial Chromosome (BAC) vector clones carrying human DNA were chosen as the intermediate templates for the sequencing of the human genome due to their inherent stability and fidelity to the genome sequence from which they were derived. In this unit, we describe a set of protocols for BAC-based array comparative genomic hybridization (aCGH) that comprise the generation of targets for printing solutions onto glass slides, the subsequent hybridization steps, and CGH analysis of a test sample compared to a reference normal sample. The BAC clones through their sequence allow the extent and gene content of numerical aberrations to be delineated by aCGH, and also provide cytogeneticists with tools for subsequent validation or fine mapping studies.

The tumour microenvironment can be a potent carcinogen, not only by facilitating cancer progression and activating dormant cancer cells, but also by stimulating tumour formation. We have previously investigated stromelysin-1/matrix metalloproteinase-3 (MMP-3), a stromal enzyme upregulated in many breast tumours, and found that MMP-3 can cause epithelial-mesenchymal transition (EMT) and malignant transformation in cultured cells, and genomically unstable mammary carcinomas in transgenic mice. Here we explain the molecular pathways by which MMP-3 exerts these effects: exposure of mouse mammary epithelial cells to MMP-3 induces the expression of an alternatively spliced form of Rac1, which causes an increase in cellular reactive oxygen species (ROS). The ROS stimulate the expression of the transcription factor Snail and EMT, and cause oxidative damage to DNA and genomic instability. These findings identify a previously undescribed pathway in which a component of the breast tumour microenvironment alters cellular structure in culture and tissue structure in vivo, leading to malignant transformation.