Epigenetic analyses in blood cells of men suspected of prostate cancer predict the outcome of biopsy better than serum PSA levels.

Lymphocytes from the peripheral blood of patients with prostate cancer-the most frequent (noncutaneous) tumor in men-display epigenetic aberrations (altered modes of allelic replication) characteristic of the malignant phenotype. The present study aims to determine whether replication aberrations add certainty to the suspicion of prostate cancer provided by the prostate-specific antigen (PSA) blood test. The allelic replication mode (whether synchronous or asynchronous) was exemplified for RB1 and AML1. These two genes normally exhibit a synchronous mode of allelic replication. Fluorescence in situ hybridization (FISH) replication assay was used for replication analyses. The FISH assays were applied to PHA-stimulated lymphocytes, established from peripheral blood samples of 35 men referred to biopsy due to suspected prostate cancer. Following biopsy 13 out of these 35 men were found positive for prostate malignancy. The FISH assay-showing asynchronous or synchronous RB1 and AML1 replication-was able to predict, respectively, the results of all biopsy-positive men and in 18 out of the 22 biopsy-negative ones. These measurements, distinguishing biopsy-positive from biopsy-negative men, were highly significant (P < 10(-8); 100% sensitivity and 81.8% specificity). Yet, distinguishing between the two groups of men based on the PSA measurements was nonsignificant (P > 0.70). The FISH replication assay applied to peripheral blood lymphocytes of 35 men referred for biopsy significantly predicted the outcome of the pathological examination, more precisely than the serum PSA test. As such, the epigenetic alteration offers a potential noninvasive blood marker, complementary to the PSA, for a preliminary prostate cancer diagnosis.

Replication timing aberrations and aneuploidy in peripheral blood lymphocytes of breast cancer patients.

BACKGROUND: Peripheral blood lymphocytes of patients with hematological malignancies or solid tumors, such as renal cell carcinoma or prostate cancer, display epigenetic aberrations (loss of synchronous replication of allelic counterparts) and genetic changes (aneuploidy) characteristic of the cancerous phenotype. This study sought to determine whether such alterations could differentiate breast cancer patients from cancer-free subjects. METHODS: The HER2 locus-an oncogene assigned to chromosome 17 whose amplification is associated with breast cancer (BCA)-and the pericentromeric satellite sequence of chromosome 17 (CEN17) were used for replication timing assessments. Aneuploidy was monitored by enumerating the copy numbers of chromosome 17. Replication timing and aneuploidy were detected cytogenetically using fluorescence in situ hybridization technology applied to phytohemagglutinin-stimulated lymphocytes of 20 women with BCA and 10 control subjects. RESULTS: We showed that both the HER2 and CEN17 loci in the stimulated BCA lymphocytes altered their characteristic pattern of synchronous replication and exhibited asynchronicity. In addition, there was an increase in chromosome 17 aneuploidy. The frequency of cells displaying asynchronous replication in the patients' samples was significantly higher (P < 10(-12) for HER2 and P < 10(-6) for CEN17) than the corresponding values in the control samples. Similarly, aneuploidy in patients' cells was significantly higher (P < 10(-9)) than that in the controls. CONCLUSIONS: The HER2 and CEN17 aberrant replication differentiated clearly between BCA patients and control subjects. Thus, monitoring the replication of these genes offers potential blood markers for the detection and monitoring of breast cancer.

The aberrant asynchronous replication - characterizing lymphocytes of cancer patients - is erased following stem cell transplantation.

BACKGROUND: Aberrations of allelic replication timing are epigenetic markers observed in peripheral blood cells of cancer patients. The aberrant markers are non-cancer-type-specific and are accompanied by increased levels of sporadic aneuploidy. The study aimed at following the epigenetic markers and aneuploidy levels in cells of patients with haematological malignancies from diagnosis to full remission, as achieved by allogeneic stem cell transplantation (alloSCT). METHODS: TP53 (a tumor suppressor gene assigned to chromosome 17), AML1 (a gene assigned to chromosome 21 and involved in the leukaemia-abundant 8;21 translocation) and the pericentomeric satellite sequence of chromosome 17 (CEN17) were used for replication timing assessments. Aneuploidy was monitored by enumerating the copy numbers of chromosomes 17 and 21. Replication timing and aneuploidy were detected cytogenetically using fluorescence in situ hybridization (FISH) technology applied to phytohemagglutinin (PHA)-stimulated lymphocytes. RESULTS: We show that aberrant epigenetic markers are detected in patients with hematological malignancies from the time of diagnosis through to when they are scheduled to undergo alloSCT. These aberrations are unaffected by the clinical status of the disease and are displayed both during accelerated stages as well as in remission. Yet, these markers are eradicated completely following stem cell transplantation. In contrast, the increased levels of aneuploidy (irreversible genetic alterations) displayed in blood lymphocytes at various stages of disease are not eliminated following transplantation. However, they do not elevate and remain unchanged (stable state). A demethylating anti-cancer drug, 5-azacytidine, applied in vitro to lymphocytes of patients prior to transplantation mimics the effect of transplantation: the epigenetic aberrations disappear while aneuploidy stays unchanged. CONCLUSIONS: The reversible nature of the replication aberrations may serve as potential epigenetic blood markers for evaluating the success of transplant or other treatments and for long-term follow up of the patients who have overcome a hematological malignancy.