Advancing a clinically relevant perspective of the clonal nature of cancer

Abstract

Cancers frequently arise as a result of an acquired genomic instability and the subsequent clonal evolution of neoplastic cells with variable patterns of genetic aberrations. Thus, the presence and behaviors of distinct clonal populations in each patient's tumor may underlie multiple clinical phenotypes in cancers. We applied DNA content-based flow sorting to identify and isolate the nuclei of clonal populations from tumor biopsies, which was coupled with array CGH and targeted resequencing. The results produced high-definition genomic profiles of clonal populations from 40 pancreatic adenocarcinomas and a set of prostate adenocarcinomas, including serial biopsies from a patient who progressed to androgen-independent metastatic disease. The genomes of clonal populations were found to have patient-specific aberrations of clinical relevance. Furthermore, we identified genomic aberrations specific to therapeutically responsive and resistant clones arising during the evolution of androgen-independent metastatic prostate adenocarcinoma. We also distinguished divergent clonal populations within single biopsies and mapped aberrations in multiple aneuploid populations arising in primary and metastatic pancreatic adenocarcinoma. We propose that our high-definition analyses of the genomes of distinct clonal populations of cancer cells in patients in vivo can help guide diagnoses and tailor approaches to personalized treatment.

Fig. 1.

Clonal analyses of a PA genome (biopsy 4050489). (A) DAPI-based DNA content analysis detected a 2.3N clonal population. The diploid and aneuploid populations were sorted for subsequent aCGH studies. (B) Multiple focal amplicons were detected only on chromosome 3 in the 2.3N population. The diploid population had a nonaberrant genome. (C) Close-up view of FANCD2 and PIK3CA amplicons in the 2.3N clonal population. Black horizontal bars represent the cores of the amplicons. Genes included in the FANCD2 amplicon cores were the following: OGG1, CAMK1, TADA3L, ARPC4, TTLL3, RPUSD3, CIDEC, JAGN1, IL17RE, FANCD2, VHL, IRAK2, TATDN2, GHRL, SEC13. Genes included in the PIK3CA amplicon core: PIK3CA, KCNMB3.

Fig. 2.

Clonal analysis of a PC metastasis initially diagnosed as adrenal cortical carcinoma. (A) DAPI-based DNA content analysis detected a 3.8N tumor population. (B) Examples of profiled chromosomes. The homozygous deletion on chromosome 10 and the interstitial deletion at 21q22 were detectable only in the flow-sorted population. (C) Locus-specific views of genomic aberrations in the 3.8N clonal population: concurrent homozygous deletions (PTEN, TP53), interstitial deletion targeting the ERG and the TMPRSS2 genes, and the amplification of the AR gene.

Fig. 3.

Genomic profiling of distinct clonal populations within a castration-resistant PC. (A) DAPI-based DNA content analysis detected two distinct tumor populations: a diploid (2N) population and an aneuploid (2.5N) population. (B) Summary of the genomic aberrations present in the two clonal populations including homozygous deletions unique to the 2.5N population. Each population showed a series of shared (black) and unique (red) genomic aberrations. (C) The aneuploid population (dark gray line) harbors additional homozygous deletions, such as the deletion of the NEDD9 gene (6p25-24). (D) Both populations show the identical focal 9p21 deletion. (E) Both populations show the identical Xq12 amplification harboring only the AR gene.

Fig. 4.

Clonal analysis of a PA rapid autopsy. DAPI-based DNA content analyses of the four sites detected a 4.5N clonal population in the pancreas, the liver, and the diaphragm, but not in the lung. The lung metastasis consisted of a 6.0N clonal population. Genomic aberrations found in the distinct aneuploid populations are listed at the right. Aberrations in black are shared by all aneuploid populations, aberrations in red are specific for one organ site, and aberrations in green were found in all populations in addition to the liver.

Fig. 5.

Clonal analyses of a multibiopsy series from a patient with PC. (A) Multiple populations were detected by DNA content flow cytometry over an 8-y time span during the clinical history of a prostate patient. Each population showed a series of shared (black) and unique (red) genomic aberrations. (B–F) Histological (HE) and FISH analysis (small box) (centromere 7, green; centromere 3, red) of the detected populations. (B) Diploid population from 2000. (C) Aneuploid population from 2000. (D) Diploid population from 2007. (E) Aneuploid population from 2007. (F) Unique (diploid) population from 2008.