by ecancer reporter Clare Sansom

It is now widely believed that adenocarcinoma of the lung in smokers arises through a gradual accumulation of mutations and genetic damage as the cells progress to malignity.

Using second-generation sequencing techniques, researchers from Genentech have completed the full DNA sequence of cells from a primary lung adenocarcinoma derived from a long-term smoker, and from normal cells from adjacent lung tissue. A comparison of the two genomes has revealed a comprehensive range of mutations in this single tumour.

In this study, following sequencing of tumour and normal tissue, DNA reads were aligned to the reference human genome sequence and larger changes, including copy number imbalance, determined using comparative genomic hybridization (CGH).

The main results showed:

• The total number of fully validated lesions in the tumour genome was extremely large, and included allelic and copy number changes, insertions, deletions and over 50,000 single nucleotide polymorphisms.

• Predicted changes in protein-coding regions of the genome, about 70% of which were validated, included 302 non-synonymous single nucleotide polymorphisms (which cause amino acid changes).

• The frequency of non-synonymous mutations was significantly lower in protein-coding genes expressed in the lung tissue than in either non-expressed genes or intergenic DNA.

• Only one observed non-synonymous mutation had already been documented: a change in the KRAS gene resulting in a substitution of the amino acid cysteine for alanine.  However, new mutations were discovered in 13 other genes that were already known to be mutated in lung adenocarcinoma.

• Many mutations, including amino-acid altering single nucleotide polymorphisms, were observed in a number of kinase genes. These included a novel mutation in the protein kinase domain of NEK9 and mutations in many genes in the EGFR-RAS-RAF-MEK-ERK pathway.

• Mutations were also observed in genes coding for RNA molecules, in pseudogenes, and in predicted transcription factor binding sites.

• 344 putative structural variations were observed, and two of these – a translocation between chromosomes 4 and 9, and a large inversion on chromosome 15 – were validated using fluorescence in situ hybridisation (FISH).

Taken together, these findings indicate that the genetic changes that arise during lung tumour development are complex and wide-ranging, and that some – such as mutations in many kinases in a single signal transduction pathway – may be redundant. Other recent whole-genome tumour studies have revealed similar trends, but few, if any, mutations have been observed in a wide range of samples. “Identification of recurrent driver mutations will require the sequencing of many more [tumour] samples”, conclude the study authors.

Reference:

Lee, W., Jiang, Z., Liu, J. and 24 others (2010). The mutation spectrum revealed by paired genome sequences from a lung cancer patient. Nature 465, 473-477.

Doi:

doi:10.1038/nature09004