Mutation screening of the TNFRSF11A gene encoding receptor activator of NF kappa B (RANK) in familial and sporadic Paget's disease of bone and osteosarcoma.

Paget's disease of bone (PDB) is a common disorder characterized by focal areas of increased and disorganized osteoclastic bone resorption, leading to bone pain, deformity, pathological fracture, and an increased risk of osteosarcoma. Genetic factors play an important role in the pathogenesis of Paget's disease. In some families, the disease has been found to be linked to a susceptibility locus on chromosome 18q21-22, which also contains the gene responsible for familial expansile osteolysis (FEO)--a rare bone dysplasia with many similarities to Paget's disease. Insertion mutations of the TNFRSF11A gene encoding Receptor Activator of NF kappa B (RANK) have recently been found to be responsible for FEO and rare cases of early onset familial Paget's disease. Loss of heterozygosity (LOH) affecting the PDB/FEO critical region has also been described in osteosarcomas suggesting that TNFRSF11A might also be involved in the development of osteosarcoma. In order to investigate the possible role of TNFRSF11A in the pathogenesis of Paget's disease and osteosarcoma, we conducted mutation screening of the TNFRSF11A gene in patients with familial and sporadic Paget's disease as well as DNA extracted from Pagetic bone lesions, an osteosarcoma arising in Pagetic bone and six osteosarcoma cell lines. No specific abnormalities of the TNFRSF11A gene were identified in a Pagetic osteosarcoma, the osteosarcoma cell lines, DNA extracted from Pagetic bone lesions, or DNA extracted from peripheral blood in patients with familial or sporadic Paget's disease including several individuals with early onset Paget's disease. These data indicate that TNFRSF11A mutations contribute neither to the vast majority of cases of sporadic or familial PDB, nor to the development of osteosarcoma.

Carcinogen-specific induction of genetic instability.

It has been proposed recently that the type of genetic instability in cancer cells reflects the selection pressures exerted by specific carcinogens. We have tested this hypothesis by treating immortal, genetically stable human cells with representative carcinogens. We found that cells resistant to the bulky-adduct-forming agent 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) exhibited a chromosomal instability (CIN), whereas cells resistant to the methylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) exhibited a microsatellite instability (MIN) associated with mismatch repair defects. Conversely, we found that cells purposely made into CIN cells are resistant to PhIP, whereas MIN cells are resistant to MNNG. These data demonstrate that exposure to specific carcinogens can indeed select for tumor cells with distinct forms of genetic instability and vice versa.

Genetic instability and darwinian selection in tumours.

Genetic instability has long been hypothesized to be a cardinal feature of cancer. Recent work has strengthened the proposal that mutational alterations conferring instability occur early during tumour formation. The ensuing genetic instability drives tumour progression by generating mutations in oncogenes and tumour-suppressor genes. These mutant genes provide cancer cells with a selective growth advantage, thereby leading to the clonal outgrowth of a tumour. Here, we discuss the role of genetic instability in tumour formation and outline future work necessary to substantiate the genetic instability hypothesis.

Characterization of MAD2B and other mitotic spindle checkpoint genes.

Aneuploidy is a characteristic of the majority of human cancers, and recent work has suggested that mitotic checkpoint defects play a role in its development. To further explore this issue, we isolated a novel human gene, MAD2B (MAD2L2), which is homologous to the spindle checkpoint gene MAD2 (MAD2L1). We determined the chromosomal localization of it and other spindle checkpoint genes, including MAD1L1, MAD2, BUB3, TTK (MPS1L1), and CDC20. In addition, we resolved the genomic intron-exon structure of the human BUB1 gene. We then searched for mutations in these genes in a panel of 19 aneuploid colorectal tumors. No new mutations were identified, suggesting that genes yet to be discovered are responsible for most of the checkpoint defects observed in aneuploid cancers.

Mutations of mitotic checkpoint genes in human cancers.

Genetic instability was one of the first characteristics to be postulated to underlie neoplasia. Such genetic instability occurs in two different forms. In a small fraction of colorectal and some other cancers, defective repair of mismatched bases results in an increased mutation rate at the nucleotide level and consequent widespread microsatellite instability. In most colorectal cancers, and probably in many other cancer types, a chromosomal instability (CIN) leading to an abnormal chromosome number (aneuploidy) is observed. The physiological and molecular bases of this pervasive abnormality are unknown. Here we show that CIN is consistently associated with the loss of function of a mitotic checkpoint. Moreover, in some cancers displaying CIN the loss of this checkpoint was associated with the mutational inactivation of a human homologue of the yeast BUB1 gene; BUB1 controls mitotic checkpoints and chromosome segregation in yeast. The normal mitotic checkpoints of cells displaying microsatellite instability become defective upon transfer of mutant hBUB1 alleles from either of two CIN cancers.