We prospectively enrolled 28 consecutive anti-HCV–negative patients with an oncohematological

disease who first underwent chemotherapy from April 2006 to November 2007. All patients were screened for hepatitis B surface antigen (HBsAg), anti-HBs (antibody to hepatitis B surface antigen), anti-HBc (antibody to hepatitis B core antigen), and anti-HCV. The diagnosis and treatment of the oncohematological diseases were based on commonly accepted criteria. For each patient, samples of plasma and PBMCs were obtained at enrollment, at months 1 and 3 during chemotherapy, and then every 3 months after treatment discontinuation. The 28 patients were treated with chemotherapy for 4-12 months Kinase Inhibitor Library in vitro and observed after its discontinuation for 6-24 months. PBMCs were isolated from 5 mL whole blood by means of Histopaque (Sigma-Aldrich, St. Louis, MO) according to a standard technique and collected in aliquots of 2 × 106 cells. The presence of HCV RNA in plasma and PBMCs of all samples collected during the study was determined as previously reported.5 The detection limit in the plasma samples was around 40 IU/mL. The sensitivity of our method to detect HCV RNA in PBMC samples was assessed using HCV-positive PBMCs diluted in PBMCs obtained from an HCV RNA–negative

patient, as described by Halfon et al.6 Briefly, 2 × 106 PBMCs from an HCV RNA–positive PLX3397 solubility dmso patient quantified at 1.8 × 104 IU/2 × 106 PBMCs was sequentially diluted (1:10) in 2 × 106 HCV RNA–negative PBMCs; in these PBMC mixtures, HCV RNA was then quantified by real-time polymerase

chain reaction. The lowest detection limit by this method was 18 IU/2 × 106 cells. As a positive control for extraction of RNA from PBMCs, glucose-6-phosphate dehydrogenase almost (G6PDH) messenger RNA was sought in all PBMC samples collected (LightCycler h-G6PDH Housekeeping Gene Set; Roche Diagnostics, Branchburg, NJ). Table 1 shows the demographic, clinical, biochemical, and serological characteristics observed at the baseline in the 28 patients enrolled (Table 1). The three HBsAg-/HBV DNA–positive patients at the baseline were treated with telbivudine or entecavir. They became HBV DNA–negative within 6 months while still under treatment and remained so throughout the observation; the 16 HBsAg-negative/anti-HBc–positive patients received lamivudine prophylaxis and never showed circulating HBsAg or HBV DNA. No plasma or PBMC sample collected during the study was HCV RNA–positive. All PBMC samples collected were positive for G6PDH messenger RNA. No patient in the present study became positive for HCV RNA in plasma or PBMCs while under chemotherapy for an oncohematological disease.