Effectiveness of nanofiltration in removing small non-enveloped viruses from three different plasma-derived products.

The objective of this study was to assess the ability of nanofiltration of albumin solution, prothrombin complex (PTC) and factor IX (FIX) to remove two small, non-enveloped DNA viruses, parvovirus B19 (B19V) and torque teno virus (TTV). Virus removal was investigated with down-scale experiments performed with sequential steps of 35-nm and 15-nm nanofiltrations of products spiked with virus DNA-positive sera. Viral loads were determined by real-time PCRs. The 15-nm nanofiltration removed more than 4.0 B19V log from all the products, TTV was reduced of more than 3.0 log from albumin solution and FIX by 35-nm and 15-nm nanofiltrations, respectively, being viral DNA undetectable after these treatments. Traces of TTV were still found in PTC after the 15-nm nanofiltration. In conclusion, nanofiltration can be efficacious in removing small naked viruses but, since viruses with similar features can differently respond to the treatment, a careful monitoring of large-scale nanofiltration should be performed.

Horse antilymphocytic globulin in hepatitis B exacerbation after bone marrow transplantation adoptive immunity transfer.

We describe the case of a HBsAg+, HBeAg+ carrier, treated with lamivudine, who experienced exacerbation of hepatitis after BMT from an anti-HBs+, anti-HBc+, anti-HBe+ donor. The serological profile of the donor and the timing of exacerbation suggested that the adoptive immunity transfer played a major pathogenetic role. Antilymphocyte globulin administration resulted in resolution of hepatitis and seroconversion to anti-HBs+. Therapy aimed at blocking the effector arm of liver damage could represent a novel approach to avoid the risk of progression to fulminant hepatitis without hampering the chances of recovery from hepatitis B.

Stretch-activated cation channels of leech neurons exhibit two activity modes.

Single-channel recordings were used to characterize two activity modes of stretch activated channels (SACs) in identified neurons of the leech. Clear-cut differences in the activity pattern of SACs from freshly desheathed cell bodies and from cultured AP cells were observed. SACs of inside-out patches, made by 'gentle' sealing and excised from cell bodies of freshly desheathed ganglia exhibited spike-like (SL) activity, with a mean channel open time (MCOT) shorter than 10 ms. Fitting of dwell open-time distributions revealed time constants shorter than 2 and 10 ms. This activity was characterized by a chord conductance of about 115 pS. SACs from cultured cells often displayed activity just after excision. MCOT exceeded 200 ms and the time constants of open-time interval distributions were longer than 10 and 100 ms. Furthermore, this activity pattern was characterized by both sub- (about 80 and 40 pS) and super-conductance (150 pS) levels, hence denoted as multiconductance (MC) mode. The percentage of open time spent at the main subconductance level (80 pS) was significantly higher in patches isolated from growth cones than in those from cell bodies of cultured neurons. The two activity modes (SL and MC) should belong to the same channel because both modes have a common main conductance value and exhibit outward rectification, stretch sensitivity and blockage by Gd3+ and gentamicin. Cytochalasin D applied to the cytoplasmic side induced activation of SACs or increased their ongoing activity. Thus, the observed differences in the expression of the two activity modes of SACs might be associated with different arrangements of the cortical cytoskeleton.

Voltage-induced activation of mechanosensitive cation channels of leech neurons.

The voltage dependence of stretch-activated cation channels in leech central neurons was studied in cell-free configurations of the patch-clamp technique. We established that stretch-activated channels excised from identified cell bodies of desheathed ganglia, as well as from neurons in culture, were slowly and reversibly activated by depolarizing membrane potentials. Negative pressure stimuli, applied to the patch pipette during a slow periodical modulation of membrane potential, enhanced channel activity, whereas positive pressures depressed it. Voltage-induced channel activation was observed, with soft glass pipettes, both in inside-out and outside-out membrane patches, at negative and positive reference potentials, respectively. The results presented in this study demonstrate that membrane depolarization induces slow activation of stretch-activated channels of leech central neurons. This phenomenon is similar to that found in Xenopus oocytes, however, some peculiar features of the voltage dependence in leech stretch-activated channels indicate that specific membrane-glass interactions might not necessarily be involved. Moreover, following depolarization, stretch-activated channels in membrane patches from neurons in culture exhibited significantly shorter delay to activation (sec) than their counterparts from neurons of freshly isolated ganglia (hundreds of sec).

Mobilization, collection, and characterization of peripheral blood hemopoietic progenitors after chemotherapy with epirubicin, paclitaxel, and granulocyte-colony stimulating factor administered to patients with metastatic breast carcinoma.

BACKGROUND: As single agents, both paclitaxel and epirubicin in combination with cytokines can mobilize peripheral blood progenitor cells (PBPCs). The authors have demonstrated previously that the combination of epirubicin and paclitaxel is very active against metastatic breast carcinoma and tolerated by patients. METHODS: Twenty-one patients with metastatic breast carcinoma received epirubicin 90 mg/m2 in combination with paclitaxel 200 mg/m2 given as a 3-hour infusion, and granulocyte-colony stimulating factor (G-CSF) starting 24 hours after chemotherapy to mobilize PBPCs. An immunophenotypic analysis for CD3, CD4, CD8, CD 19, CD33, CD34, and CD38 antigen expression was performed on apheresis products. Eighteen patients underwent high dose chemotherapy and were engrafted with PBPCs primed with paclitaxel, epirubicin, and G-CSF. RESULTS: The median number of circulating CD34+ cells at peak was 70/microL; in the patients less heavily pretreated, it was 106.7/microL. The mean number of CD34+, CD34+/CD33-, and CD34+/CD38- cells/kg collected per apheresis was 6.3 x 10(6), 2.0 x 10(6), and 0.18 x 10(6), respectively. The mean number of CD34+ cells/kg per apheresis was 7.8 x 10(6) when the preleukapheresis CD34+ cell count was more than 50/microL and 0.9 x 10(6) when the CD34+ cell count was less than 50/microL. The mean number of CD3+, CD4+, and CD8+ cells/kg collected per apheresis was 90 x 10(6), 50 x 10(6), and 30 x 10(6), respectively. CONCLUSIONS: Epirubicin plus paclitaxel in combination with G-CSF mobilizes PBPCs, including more primitive progenitors capable of supporting myeloablative treatment. Moreover, the mononuclear cells collected in this study contained high levels of cytotoxic effector cells suitable for ex vivo manipulation to augment the antitumor effect.

High-dose thiotepa and melphalan with hemopoietic progenitor support following induction therapy with epirubicin-paclitaxel-containing regimens in metastatic breast cancer (MBC).

BACKGROUND: Preliminary data from phase III randomized studies have failed to show benefit of HDC given as consolidation after anthracycline and alkylating-based chemotherapy in metastatic breast cancer (MBC). Moderate activity of induction regimens and selection of chemoresistant clones are among the possible reasons for these disappointing results. We therefore have designed a phase II study where high-dose alkylating agents are given as consolidation after an induction treatment including the most active agents (epirubicin and paclitaxel) without alkylating agents. PATIENTS AND METHODS: Patients with MBC not previously treated with chemotherapy for metastatic disease were eligible. After six courses of epirubicin-paclitaxel +/- gemcitabine patients received a course of thiotepa 600 mg/m2 + melphalan 160 mg/m2 with hemopoietic support. Pharmacokinetic parameters of thiotepa and melphalan were measured and related to treatment outcomes. The L-VEF of the patients was monitored before and after treatment. RESULTS: Forty-eight patients have been treated. Before HDC 14 patients were in CR, and 34 in PR. A median of 6.92 x 10(6) (range 1.53-16.6) CD34+ cells/kg were reinfused after HDC. Median days (range) to neutrophils > 0.5 x 10(9)/l and platelets > 20,000 x 10(9)/l were 9.5 (9-33) and 10 days (9-32), respectively. Symptomatic CHF was observed in two patients (4.1%). Cmax and AUC of thiotepa showed a linear relationship with time to progression (TTP) and overall survival (OS): r2 = 0.6. After HDC the conversion rate from PR to CR was 44.1%. At five years progression-free and overall survival rates are 37.5% and 65%, respectively. A treatment-related death was observed. CONCLUSIONS: High-dose thiotepa and melphalan after an epirubicin-paclitaxel-containing treatment is feasible, devoid of significant cardiotoxicity and very active. Pharmacokinetic parameters of high-dose thiotepa might be linked to treatment outcome.

Variable correlation between 6-mercaptopurine metabolites in erythrocytes and hematologic toxicity: implications for drug monitoring in children with acute lymphoblastic leukemia.

Nineteen pediatric patients affected by acute lymphoblastic leukemia (ALL) were examined weekly with respect to 6-mercaptopurine nucleotide (6-MPN) and 6-thioguanine nucleotide (6-TGN) levels in erythrocytes during the course of maintenance treatment with 6-MP 50 mg/m2 per d and results were related to various parameters of bone marrow function to assess, in the same individual, the level of reliability of 6-MP metabolites in predicting a later change in peripheral blood cell counts. Median values for 6-MPN and 6-TGN were 57 and 200 pmol/8 x 10(8) erythrocytes, respectively, as measured by reversed-phase high-performance liquid chromatography (HPLC). 6-TGN levels in erythrocytes were inversely related with white blood cell count (r = -0.463, p < 0.0001, n = 361), absolute neutrophil count (r = -0.386, p < 0.0001, n = 347), erythrocyte (r = -0.354, p < 0.0001, n = 287), and platelet counts (r = -0.24, p < 0.0001, n = 319) in the majority of patients (n = 10-12), while no correlation was found for 6-MPN. In the remaining children, no evidence of correlation was demonstrated between 6-TGN levels and myelotoxicity. The results confirm the role of 6-TGN as the reference cytotoxic metabolite for evaluating the exposure to 6-MP and identifying treatment compliance in ALL children but indicate the limits of a follow-up based solely on metabolite levels and suggest that a more correct approach remains the double monitoring of 6-TGN and blood cell count with differential.