Genomic alterations indicate tumor origin and varied metastatic potential of disseminated cells from prostate cancer patients

Abstract

Disseminated epithelial cells can be isolated from the bone marrow of a far greater fraction of prostate-cancer patients than the fraction of patients who progress to metastatic disease. To provide a better understanding of these cells, we have characterized their genomic alterations. We first present an array comparative genomic hybridization method capable of detecting genomic changes in the small number of disseminated cells (10-20) that can typically be obtained from bone marrow aspirates of prostate-cancer patients. We show multiple regions of copy-number change, including alterations common in prostate cancer, such as 8p loss, 8q gain, and gain encompassing the androgen-receptor gene on Xq, in the disseminated cell pools from 11 metastatic patients. We found fewer and less striking genomic alterations in the 48 pools of disseminated cells from patients with organ-confined disease. However, we identify changes shared by these samples with their corresponding primary tumors and prostate-cancer alterations reported in the literature, evidence that these cells, like those in advanced disease, are disseminated tumor cells (DTC). We also show that DTCs from patients with advanced and localized disease share several abnormalities, including losses containing cell-adhesion genes and alterations reported to associate with progressive disease. These shared alterations might confer the capability to disseminate or establish secondary disease. Overall, the spectrum of genomic deviations is evidence for metastatic capacity in advanced-disease DTCs and for variation in that capacity in DTCs from localized disease. Our analysis lays the foundation for elucidation of the relationship between DTC genomic alterations and progressive prostate cancer.

Figure 1

Reproducibility of array CGH on RCGA-amplified samples from 10-20 cells. Three pools of 20 AdvDCs were collected independently and amplified independently. Arrays for the three biological replicates were run in parallel against the same female reference. A. Scatterplots of the normalized (abbreviated “norm.”) log₂-ratio values from each possible pair of replicates. The Pearson’s correlation coefficient (r) for each pair is given. B. The top plot is a whole-genome profile of the normalized log₂-ratios for one of the replicates. The normalized average log₂ ratio for each BAC is plotted on the y-axis and the midpoint of the genomic position (Build 35) of each BAC on the array is plotted on the x-axis. The three bottom graphs are whole-genome plots of deviant segments for each of the three replicates. The log2-ratio value, as determined by CBS, of each segment exceeding chromosome-specific thresholds is plotted on the y-axis, and the genome position of the deviant segment, as defined by the encompassed BACs, is plotted on the x-axis. Chromosome numbers are indicated along the x-axis.

Figure 2

Frequency of deviations in AdvDCs (n = 11). The frequency of AdvDC samples with a deviation (y-axis) is plotted at the genomic position of the midpoint of each BAC encompassed by the deviation. Blue presents the frequency of gains, and red represents the frequency of losses.

Figure 3

Comparison of copy-number changes observed in LocDC samples, matching primary tumors and AdvDC samples. A. Percent of deviations observed across the genome in LocDCs (dots) overlaid on the frequencies for primary tumors (bars). B. Percent of deviations observed across the genome in LocDCs (dots) overlaid on the frequencies for AdvDCs (bars). Light blue and orange dots in both A and B indicate the percentage of LocDC samples (n = 48) with observed copy-number change encompassing each BAC on the array. Blue and red bars give the frequency of copy-number change in the nine matching primary tumor samples (A) or in the 11 AdvDC samples (B). Deviation frequencies of zero are not plotted. The x-axis is the midpoint of the genomic position of each BAC on the array, with chromosome numbers indicated. Note that copy-number loss of chromosome X and gain of chromosome Y represent the normal state for comparisons of male test and female reference genomic DNAs.