Aneuploidy arises at early stages of Apc-driven intestinal tumorigenesis and pinpoints conserved chromosomal loci of allelic imbalance between mouse and human

Abstract

Although chromosomal instability characterizes the majority of human colorectal cancers, the contribution of genes such as adenomatous polyposis coli (APC), KRAS, and p53 to this form of genetic instability is still under debate. Here, we have assessed chromosomal imbalances in tumors from mouse models of intestinal cancer, namely Apc(+/1638N), Apc(+/1638N)/KRAS(V12G), and Apc(+/1638N)/Tp53-/-, by array comparative genomic hybridization. All intestinal adenomas from Apc(+/1638N) mice displayed chromosomal alterations, thus confirming the presence of a chromosomal instability defect at early stages of the adenoma-carcinoma sequence. Moreover, loss of the Tp53 tumor suppressor gene, but not KRAS oncogenic activation, results in an increase of gains and losses of whole chromosomes in the Apc-mutant genetic background. Comparative analysis of the overall genomic alterations found in mouse intestinal tumors allowed us to identify a subset of loci syntenic with human chromosomal regions (eg, 1p34-p36, 12q24, 9q34, and 22q) frequently gained or lost in familial adenomas and sporadic colorectal cancers. The latter indicate that, during intestinal tumor development, the genetic mechanisms and the underlying functional defects are conserved across species. Hence, our array comparative genomic hybridization analysis of Apc-mutant intestinal tumors allows the definition of minimal aneuploidy regions conserved between mouse and human and likely to encompass rate-limiting genes for intestinal tumor initiation and progression.

Figure 1

Heat map visualization of the CGH results for the three groups of tumors analyzed in the present study. After normalization and two-log smoothing of the array CGH ratios from the 30 tumor samples and four normal intestinal epithelia, the data were loaded onto SpotFire Decision Site 8.1 to obtain the heat map here represented. Data are ordered in the X bar according to the sample genotypes as normal (N), Apc^+/1638N (Apc), Apc^1638N/KRAS^V12G (Ras), and Apc^+/1638N/Tp53^−/− (p53), and in the Y bar according to the chromosome and Mb position of the BAC clones (color code at both sides of the heat map). The color code of the samples indicates BAC copy number changes: green, loss; red, gain; and black, no change.

Figure 2

LOH analysis performed on Apc^+/1638N F1 C57BL/6J × Ola/129 mouse tumors by direct nucleotide sequencing of single nucleotide polymorphisms (SNPs) validates the array CGH results. The examples are relative to three SNPs on chromosome 5: rs3704966, rs4136991, and rs4136994. A: Nucleotide sequence analysis of SNP rs3704966 showing retention of both alleles in the tumor sample. B and C: Nucleotide sequence analysis of SNPs rs3704966 or rs4136991 and rs4136994, respectively, each showing hetero- and hemizygosity in the normal and tumor DNA samples, respectively. The arrows indicate the polymorphic nucleotide position for each SNP sequence.

Figure 3

Figure 3. Overview of the chromosome copy number changes throughout the 30 mouse intestinal tumors analyzed in this study. Total numbers of all significant gains or loss regions found to be affected by copy number alterations in more than or equal to two tumor samples from at least two independent groups have been plotted separately according to the tumor genotype. Complete or partial chromosome gains and losses are shown separately. Regions are ordered according to their mapped positions along the chromosomes. G, gain; L, loss of the corresponding regions.