Application of a modified real-time PCR technique for relative gene copy number quantification to the determination of the relationship between NKX3.1 loss and MYC gain in prostate cancer.

in granulocytes is ϳ2.4-fold greater than that in mononuclear cells (Table 1). The clinical significance of these results will also likely be in the realm of heterozy-gosity testing. In addition, some individuals with ␣-idu-ronidase deficiency have undergone bone marrow or cord blood transplantation therapy (22, 27), and here, too, biochemical monitoring of enzyme activities in mixed leukocyte preparations could at times provide misleading results, suggesting the possibility of loss of engraftment when, instead, the distribution of types of leukocytes in the peripheral blood has changed. In summary, this study provides data regarding six lysosomal enzymes that are frequently measured and shows that all six enzymes have significantly different activities per gram of cell protein in mononuclear cells compared with granulocytes. These data have clinical implications in the areas of biochemical diagnosis of patients having partial deficiencies of lysosomal enzyme activities, in heterozygosity testing, and in the monitoring of patients who have undergone bone marrow or cord blood transplantation. Unusual thermolability properties of ␤-hexosaminidase: studies of enzyme from cultured cells and clinical implications. Acid hydrolases in leukocytes and platelets of normal subjects and in patients with Gaucher's and Fabry's disease.ties of lysosomal hydrolases in fractionated populations of human peripheral blood cells. A and ␤-galac-tosidase activities in leukocytes and lymphocytes from normal and psychiatric subjects. et al. Cord-blood transplants from unrelated donors in patients with Hurler's syndrome. Gene amplifications and deletions play an important role in the pathogenesis of solid tumors, including prostate cancer. Real-time PCR is a powerful tool for quantitative DNA analysis, particularly when starting quantities of tumor tissue are minimal (1–5). In the present study, we describe a modification of the 2 Ϫ⌬⌬CT method, which recently was shown to be suitable for relative gene expression analyses (6). We used our technique to analyze prostate cancer cell lines and tissue samples to determine the relationship between homeodomain-containing transcription factor 3.1 (NKX3.1) and MYC gene copy number alterations in this tumor type. The specimens analyzed included (a) blood samples from healthy donors; (b) the prostate cancer cell lines DU145 (ATCC no. HTB-81) and PC3 (ATCC no. CRL-1435) and their derived sublines DU145MN1, PC3-N, and PC3-125-1L (7); (c) the colorectal cell line COLO320DM (ATCC no. CCL-220), which harbors a high-level MYC amplification; and (d) primary prostate adenocarcinoma samples obtained after radical prostatectomy from previously untreated patients. The specimens were histologi