Anti-Hepatitis B Virus Activity and Metabolism of 2′,3′-Dideoxy-2′,3′-Didehydro-β-l(?)-5-Fluorocytidine

2′,3′-Dideoxy-2′,3′-didehydro-β-l(−)-5-fluorocytidine [l(−)Fd4C] was found to be at least 10 times more potent than β-l-2′,3′-dideoxy-3′-thiacytidine [l(−)SddC; also called 3TC, or lamivudine]against hepatitis B virus (HBV) in culture. Its cytotoxicity against HepG2 growth in culture was also greater than that of l(−)SddC (3TC). There was no activity of this compound against mitochondrial DNA synthesis in cells at concentrations up to 10 μM. The dynamics of recovery of virus from the medium of cells pretreated with equal drug concentrations were slower with l(−)Fd4C than with l(−)SddC (3TC). l(−)Fd4C could be metabolized to mono-, di-, and triphosphate forms. The degree of l(−)Fd4C phosphorylation to the 5′-triphosphate metabolite was higher than the degree of l(−)SddC (3TC) phosphorylation when equal extracellular concentrations of the two drugs were used. The apparent K_m of l(−)Fd4C phosphorylated metabolites formed intracellularly was higher than that for l(−)SddC (3TC). This may be due in part to a difference in the behavior of l(−)Fd4C and l(−)SddC (3TC) towards cytosolic deoxycytidine kinase. Furthermore, l(−)Fd4C 5′-triphosphate was retained longer within cells than l(−)SddC (3TC) 5′-triphosphate. l(−)Fd4C 5′-triphosphate inhibited HBV DNA polymerase in competition with dCTP with a K_i of 0.069 ± 0.015 μM. Given the antiviral potency and unique pharmacodynamic properties of l(−)Fd4C, this compound should be considered for development as an expanded-spectrum anti-HBV drug.Hepatitis B virus (HBV) infection is one of the most serious health issues in the world today (1, 3). β-l(−)-2′,3′-Dideoxy-3′-thiacytidine [l(−)SddC; also called 3TC, or lamivudine] (Fig. ​(Fig.1)1) is the first β-l(−) nucleoside analog identified by us and others to have potent activity against HBV in culture (4, 8, 12, 17). This drug exerts its action by inhibiting HBV DNA synthesis due to the preferential interaction of the l(−)SddC (3TC) 5′-triphosphate metabolite with HBV DNA polymerase (4). Unlike dideoxycytidine (ddC, or zalcitabine), a β-d(+) nucleoside analog with the natural nucleoside conformation in DNA and RNA, l(−)SddC (3TC) does not have potent activity against mitochondrial DNA (mtDNA) synthesis, which is important for maintaining the function of cells (4). Clinical trials of l(−)SddC (3TC) for the treatment of chronic HBV infection are ongoing and look promising (2, 7, 13, 16, 18, 19). Its potential value for HBV-infected patients undergoing liver transplantation is also being evaluated, since l(−)SddC (3TC) can suppress HBV DNA serum levels in these patients. However, apparent l(−)SddC (3TC)-resistant HBV emerged in some patients upon long-term treatment (13, 16, 18). The HBV-resistant genotype appears to be associated with the mutation of methionine to valine or isoleucine in the YMDD motif of HBV DNA polymerase (13, 16, 18). This mutation was previously demonstrated and could render duck HBV resistant to l(−)SddC (3TC) (11). It is not clear if this mutation alone can lead to resistance and, if so, to what degree HBV resistance to l(−)SddC (3TC) develops. Open in a separate windowFIG. 1Structures of l(−)deoxycytidine analogs.One approach to overcoming clinical drug resistance is to use higher dosages of l(−)SddC (3TC) given the therapeutic index of the compound in vitro. However, the potency of l(−)SddC (3TC) against HBV in the clinic could be a limiting factor given the dosage application. The antiviral potency is determined not only by its antiviral activity but also by the pharmacodynamic nature of its active metabolite, l(−)SddC (3TC) 5′-triphosphate, in vivo. A more potent compound with more favorable pharmacodynamic behavior of intracellularly active metabolites would be worth exploring.In the studies reported herein, we describe the anti-HBV activity, metabolism, and pharmacodynamic properties of 2′,3′-dideoxy-2′,3′-didehydro-β-l(−)-5-fluorocytidine [l(−)Fd4C](Fig. 1) in comparison with those of l(−)SddC (3TC), including its behavior toward deoxycytidine kinase and the interaction of l(−)Fd4C 5′-triphosphate with virion-associated HBV DNA polymerase. Preliminary studies of the anti-HBV and anti-human immunodeficiency virus (HIV) activities of l(−)Fd4C were previously reported by us and others (10, 15).     

In Vitro Activity of Ceftolozane Alone and in Combination with Tazobactam against Extended-Spectrum-β-Lactamase-Harboring Enterobacteriaceae

Ceftolozane, formally CXA-101, is a new antipseudomonal cephalosporin that is also active in vitro against Enterobacteriaceae but is vulnerable to extended-spectrum β-lactamases (ESBLs). The addition of tazobactam is intended to broaden coverage to most ESBL-producing Escherichia coli and Klebsiella pneumonia as well as other Enterobacteriaceae. The in vitro activities of ceftolozane-tazobactam combinations against 67 clinically and molecularly characterized ESBL-producing isolates were examined by checkerboard MIC testing to evaluate their potential clinical feasibility and to assess the optimal tazobactam concentrations to be used in MIC determinations of ceftolozane. Isolates included those from E. coli (n = 32), K. pneumoniae (n = 19), Enterobacter cloacae (n = 15), and Citrobacter freundii (n = 1). Checkerboard experiments were performed to study interactions over the range of 0.008 to 64 mg/liter ceftolozane and 0.063 to 32 mg/liter tazobactam using 2-fold-dilution series. The MIC₅₀ and MIC₉₀ of ceftolozane alone for all isolates were 16 and ≥64 mg/liter, respectively. Increasing concentrations of tazobactam resulted in decreasing MICs of ceftolozane. The 50th and 90th percentile concentrations of tazobactam required to reduce the MIC of ceftolozane to 8 mg/liter for all organisms in this ESBL collection were 0.5 and 4 mg/liter, respectively. For E. coli, K. pneumoniae, and E. cloacae, these values were 0.5 and 2, 1 and 16, and 0.5 and 4 mg/liter, respectively. When combined with a fixed amount of 4 mg/liter tazobactam (current CLSI concentration used for susceptibility testing), 90% of the isolates would have an MIC of ≤4 mg/liter. The combination ceftolozane-tazobactam is a promising alternative option for treating infections due to ESBL-harboring isolates.     

Cellular Pharmacology of the Anti-Hepatitis B Virus Agent β-l-2′,3′-Didehydro-2′,3′-Dideoxy-N4-Hydroxycytidine: Relevance for Activation in HepG2 Cells

ß-l-2′,3′-Didehydro-2′,3′-dideoxy-N⁴-hydroxycytidine (l-Hyd4C) was demonstrated to be an effective and highly selective inhibitor of hepatitis B virus (HBV) replication in HepG2.2.15 cells (50% effective dose [ED₅₀] = 0.03 μM; 50% cytotoxic dose [CD₅₀] = 2,500 μM). In the present study, we investigated the intracellular pharmacology of tritiated l-Hyd4C in HepG2 cells. l-[³H]Hyd4C was shown to be phosphorylated extensively and rapidly to the 5′-mono-, 5′-di-, and 5′-triphosphate derivatives. Other metabolites deriving from a reduction or removal of the NHOH group of l-Hyd4C could not be detected, although both reactions were described as the primary catabolic pathways of the stereoisomer ß-d-N⁴-hydroxycytidine in HepG2 cells. Also, the formation of liponucleotide metabolites, such as the 5′-diphosphocholine derivative of l-Hyd4C, as described for some l-deoxycytidine analogues, seems to be unlikely. After incubation of HepG2 cells with 10 μM l-[³H]Hyd4C for 24 h, the 5′-triphosphate accumulated to 19.4 ± 2.7 pmol/10⁶ cells. The predominant peak belonged to 5-diphosphate, with 43.5 ± 4.3 pmol/10⁶ cells. The intracellular half-life of the 5′-triphosphate was estimated to be 29.7 h. This extended half-life probably reflects a generally low affinity of 5′-phosphorylated l-deoxycytidine derivatives for phosphate-degrading enzymes but may additionally be caused by an efficient rephosphorylation of the 5′-diphosphate during a drug-free incubation. The high 5′-triphosphate level and its extended half-life in HepG2 cells are consistent with the potent antiviral activity of l-Hyd4C.A large number of nucleoside analogues have been described as inhibitors of hepatitis B virus (HBV) and HIV replication. Recently l-nucleoside analogues in particular have gained increasing interest. They are characterized by an opposite configuration from that of the natural d-nucleoside analogues and represent one of the most attractive groups of antiretroviral compounds, including ß-l-2′,3′-dideoxy-3-thiacytidine (3TC) and its 5-fluoro derivative (FTC), ß-l-2′,3′-didehydro-2′,3′-dideoxy-cytidine (l-d4C) and its 5-fluoro derivative (l-d4FC), ß-l-thymidine, ß-l-fluoroarabinosylyluracil (l-FMAU), and ß-l-2′,3′-didehydro-2′,3′-dideoxy-2′-fluoro-cytidine (l-2′Fd4C) (3, 5, 22).Some of them not only have been found to be more potent than their corresponding d-nucleosides but seem to exhibit lower cytotoxicity and have been proved to be effective and selective agents for the treatment of chronic hepatitis B virus infections (4). However, only long-term therapy with a single nucleoside for several years was shown to be able to completely suppress HBV DNA in serum of patients and to reverse the progression of the disease. The disadvantage connected with such therapy regimens is the development of drug-resistant HBV strains (22). Therefore, the challenge will be to develop more-efficient drugs for shorter treatment regimens and to combine them to reach synergistic or at least additive drug action. This approach has been described not only as being highly efficient for the treatment of HIV infections but also as preventing the development of resistant mutants. Therefore, AIDS therapy is considered a model for future therapy of chronic HBV infections (17).Recently we described a series of new ß-l-N⁴-hydroxydeoxycytidine and ß-l-5-methyl-deoxycytidine derivatives as inhibitors of HBV replication. Between them, ß-l-2′,3′-didehydro-2′,3′-dideoxy-N⁴-hydroxycytidine (l-Hyd4C) (Fig. ​(Fig.1)1) emerged as the most effective in suppression of virus production in HepG2.2.15 cells (50% effective dose [ED₅₀] = 0.03 μM), displaying an extremely low cytotoxicity (50% cytotoxic dose [CD₅₀] for HepG2 cells = 2,500 μM) (12).Open in a separate windowFIG. 1.Structure of l-Hyd4C and possible metabolites formed by reduction (l-d4C) or by deamination (l-d4U).These encouraging features have prompted us to investigate the cellular pharmacology of l-Hyd4C in a hepatic cell line. This included the activation of this unnatural l-deoxycytidine nucleoside to its 5′-mono-, 5′-di-, and 5′-triphosphate, the search for other metabolites, and the estimation of the intracellular half-lives (t_1/2) of the 5′-di- and 5′-triphosphate of l-Hyd4C.(This work was presented in part at BIT''s 5th Anniversary Congress of International Drug Discovery Science and Technology, 7 to 13 November 2007, Xi''an and Beijing, China.)     

PSI-7851, a Pronucleotide of β-d-2′-Deoxy-2′-Fluoro-2′-C-Methyluridine Monophosphate,Is a Potent and Pan-Genotype Inhibitor of Hepatitis C Virus Replication

The hepatitis C virus (HCV) NS5B RNA polymerase facilitates the RNA synthesis step during the HCV replication cycle. Nucleoside analogs targeting the NS5B provide an attractive approach to treating HCV infections because of their high barrier to resistance and pan-genotype activity. PSI-7851, a pronucleotide of β-d-2′-deoxy-2′-fluoro-2′-C-methyluridine-5′-monophosphate, is a highly active nucleotide analog inhibitor of HCV for which a phase 1b multiple ascending dose study of genotype 1-infected individuals was recently completed (M. Rodriguez-Torres, E. Lawitz, S. Flach, J. M. Denning, E. Albanis, W. T. Symonds, and M. M. Berry, Abstr. 60th Annu. Meet. Am. Assoc. Study Liver Dis., abstr. LB17, 2009). The studies described here characterize the in vitro antiviral activity and cytotoxicity profile of PSI-7851. The 50% effective concentration for PSI-7851 against the genotype 1b replicon was determined to be 0.075 ± 0.050 μM (mean ± standard deviation). PSI-7851 was similarly effective against replicons derived from genotypes 1a, 1b, and 2a and the genotype 1a and 2a infectious virus systems. The active triphosphate, PSI-7409, inhibited recombinant NS5B polymerases from genotypes 1 to 4 with comparable 50% inhibitory concentrations. PSI-7851 is a specific HCV inhibitor, as it lacks antiviral activity against other closely related and unrelated viruses. PSI-7409 also lacked any significant activity against cellular DNA and RNA polymerases. No cytotoxicity, mitochondrial toxicity, or bone marrow toxicity was associated with PSI-7851 at the highest concentration tested (100 μM). Cross-resistance studies using replicon mutants conferring resistance to modified nucleoside analogs showed that PSI-7851 was less active against the S282T replicon mutant, whereas cells expressing a replicon containing the S96T/N142T mutation remained fully susceptible to PSI-7851. Clearance studies using replicon cells demonstrated that PSI-7851 was able to clear cells of HCV replicon RNA and prevent viral rebound.Hepatitis C virus (HCV) currently affects more than 170 million people worldwide. Approximately 70% of infected individuals develop chronic hepatitis, among whom about 20% will develop liver cirrhosis and fibrosis and up to 5% will progress to hepatocellular carcinoma (2). The current standard of care (SOC), which combines pegylated alpha interferon (PegIFN-α) and ribavirin (RBV), has limited efficacy in providing a sustained virological response (SVR), especially in individuals with HCV genotype 1 (∼50%), the most prevalent genotype in Western countries (8, 11, 35). The impact of genetic diversity of HCV in patients receiving SOC therapy has been reviewed (26): SVR rates are higher in patients infected with genotype 2 or 3 (∼80%), patients infected with genotype 4 appear to have a slightly better SVR rate (∼60%) than patients infected with genotype 1, and patients infected with genotypes 5 and 6 may achieve an SVR at a level between those of genotypes 1 and 2/3. In addition to the variability in efficacy, the lengthy treatment (24 to 48 weeks) with SOC is frequently associated with undesirable side effects that may include anemia, fatigue, and depression (7). There is an urgent medical need to develop anti-HCV therapies that are safer and more effective. Direct-acting antivirals (DAAs) are compounds that target a specific viral protein. Currently, four major classes of DAAs are being investigated in phase II or III clinical trials: NS3 protease inhibitors, NS5A inhibitors, allosteric nonnucleoside NS5B polymerase inhibitors, and nucleoside/-tide NS5B polymerase inhibitors (21, 27, 46). Challenges for these DAAs include safety, pan-genotypic activity, and/or emergence of resistant viruses. An effective antiviral therapy against hepatitis C should encompass a broad spectrum of activity against all HCV genotypes, shorten treatment duration, have minimal side effects, and have a high barrier to resistance.The HCV NS5B RNA-dependent RNA polymerase (Pol) is a critical component of the replicase complex and is responsible for initiating and catalyzing viral RNA synthesis (16, 32, 58). There is no human homolog of this protein, and it is absolutely required for viral infectivity (19). As a result, the HCV NS5B is an attractive target for the development of antiviral compounds. There are two major classes of NS5B inhibitors: nucleoside analogs, which are anabolized to their active triphosphates and act as alternative substrates for the polymerase, and nonnucleoside inhibitors (NNIs), which bind to allosteric regions on the protein. Two major drawbacks associated with NNIs are that the activity appears to vary significantly among different HCV genotypes and even subtypes (15, 33) and that there is a relatively low barrier for resistance as evidenced by the numerous naturally occurring resistant variants reported in the literature (18). In contrast, nucleoside analogs are similarly active across HCV genotypes (13, 15, 33) and have a higher barrier of resistance compared to the NNIs and NS3 protease inhibitors (36). To date only two amino acid changes within the NS5B polymerase that confer resistance to nucleoside inhibitors have been identified: S96T and S282T (1, 29). The S96T mutation confers resistance to 4′-azidocytidine (R1479), while the S282T mutation is resistant to a number of 2′-C-methyl-modified nucleoside inhibitors (1, 29, 38, 43).In order for nucleoside analogs to be active as alternative substrates, they must first be phosphorylated by cellular kinases to their corresponding 5′-triphosphates, which are active alternative substrate inhibitors for the NS5B polymerase. The efficiency of these metabolic steps, the stability of the triphosphates, and the affinity of the triphosphates for the NS5B polymerase are all important factors in determining the antiviral activities of nucleoside inhibitors. PSI-6130, 2′-F-2′-C-methylcytidine, was previously shown to be a specific inhibitor of HCV RNA replication in the replicon assay system (52). However, when the uridine analog, 2′-F-2′-C-methyluridine (referred to as PSI-6206), was tested in the replicon assay, it failed to inhibit HCV RNA synthesis due to the inability of cellular enzymes to metabolize PSI-6206 to its triphosphate, PSI-7409 (5, 34, 42). Biochemical studies with PSI-7409 showed that this compound was able to inhibit RNA synthesis mediated by the HCV replicase complex and by purified recombinant HCV NS5B polymerase (34, 42). Furthermore, in vitro stability studies using primary human hepatocytes demonstrated that PSI-7409 has a significantly longer half-life (t_1/2, 38 h) than PSI-6130-TP (t_1/2, 4.7 h), which could be a desirable pharmacologic benefit (34).In order to bypass the initial nonproductive phosphorylation step of PSI-6206, the phosphoramidate prodrug methodology was explored as an approach to deliver 2′-F-2′-C-methyluridine monophosphate (47, 48). An extensive series of phosphoramidate prodrugs were synthesized, and PSI-7851 demonstrated the desired characteristics with regard to activity and in vitro toxicity. Herein we present the results of in vitro studies characterizing PSI-7851, a potent and specific anti-HCV compound with pan-genotype activity.     

SILEN-C3, a Phase 2 Randomized Trial with Faldaprevir plus Pegylated Interferon α-2a and Ribavirin in Treatment-Naive Hepatitis C Virus Genotype 1-Infected Patients

Faldaprevir is an investigational hepatitis C virus (HCV) NS3/4A protease inhibitor which, when administered for 24 weeks in combination with pegylated interferon α-2a and ribavirin (PegIFN/RBV) in treatment-naive patients in a prior study (SILEN-C1; M. S. Sulkowski et al., Hepatology 57:2143–2154, 2013, doi:10.1002/hep.26276), achieved sustained virologic response (SVR) rates of 72 to 84%. The current randomized, open-label, parallel-group study compared the efficacy and safety of 12 versus 24 weeks of 120 mg faldaprevir administered once daily, combined with 24 or 48 weeks of PegIFN/RBV, in 160 treatment-naive HCV genotype 1 patients. Patients with maintained rapid virologic response (HCV RNA of <25 IU/ml at week 4 and undetectable at weeks 8 and 12) stopped all treatment at week 24, otherwise they continued PegIFN/RBV to week 48. SVR was achieved by 67% and 74% of patients in the 12-week and 24-week groups, respectively. Virologic response rates were lower in the 12-week group from weeks 2 to 12, during which both groups received identical treatment. SVR rates were similar in both groups for patients achieving undetectable HCV RNA. Most adverse events were mild or moderate, and 6% of patients in each treatment group discontinued treatment due to adverse events. Once-daily faldaprevir at 120 mg for 12 or 24 weeks with PegIFN/RBV resulted in high SVR rates, and the regimen was well tolerated. Differences in the overall SVR rates between the 12-week and 24-week groups were not statistically significant and possibly were due to IL28B genotype imbalances; IL28B genotype was not tested, as its significance was not known at the time of the study. These results supported phase 3 evaluation. (This study has been registered at ClinicalTrials.gov under registration no. {"type":"clinical-trial","attrs":{"text":"NCT00984620","term_id":"NCT00984620"}}NCT00984620).     

Tn6249, a New Tn6162 Transposon Derivative Carrying a Double-Integron Platform and Involved with Acquisition of the blaVIM-1 Metallo-β-Lactamase Gene in Pseudomonas aeruginosa

The In70.2 integron platform appears to be a conserved structure involved in the dissemination of the bla_VIM-1 metallo-β-lactamase gene in Pseudomonas aeruginosa. The genetic context of the In70.2 integron platform from P. aeruginosa VR-143/97, the VIM-1-producing index strain isolated in Italy in 1997, was fully characterized by a next-generation sequencing approach refined by conventional sequencing. The In70.2 integron platform from VR-143/97 was found to be associated with a defective Tn402-like transposon inserted into the urf2 gene of a Tn3 family transposon of an original structure, named Tn6249, which also carried a partially deleted mer operon and an In90 integron platform in a tail-to-tail orientation. Tn6249 was inserted into a PACS171b-like genomic island, which was in turn inserted into the endA gene of the Pseudomonas chromosomal backbone. Tn6249 showed a similar structure and a conserved location with respect to that of Tn6060, a Tn3 family transposon associated with In70.2 and carrying a double-integron platform, which was detected in a VIM-1-producing P. aeruginosa strain isolated in Australia in 2008. Both Tn6249 and Tn6060 are apparently derived from Tn6162, a mercury resistance transposon carrying an integron platform, which was found in P. aeruginosa isolates from different geographic locations. The conservation of the genetic context of Tn6249 and Tn6060 suggests an in situ evolution of these elements after the insertion of a Tn6162-like ancestor into the PACS171b-like genomic island (GI) present in the genome of a successful widespread P. aeruginosa clonal lineage.     

NCI-Sponsored Trial for the Evaluation of Safety and Preliminary Efficacy of 3′-Deoxy-3′-[18F]fluorothymidine (FLT) as a Marker of Proliferation in Patients with Recurrent Gliomas: Preliminary Efficacy Studies

Purpose  3′-Deoxy-3′-[¹⁸F]fluorothymidine ([¹⁸F]FLT) is being developed for imaging cellular proliferation. The goals were to explore the capacity of FLT-positron emission tomography (PET) to distinguish between recurrence and radionecrosis in gliomas and compare the results to those obtained with 2-fluoro-2-deoxy-d-glucose (FDG). Procedures  Fifteen patients with tumor recurrence and four with radionecrosis, determined by clinical course and magnetic resonance imaging results, were studied by dynamic [¹⁸F]FLT-PET with arterial blood sampling. A two-tissue compartment four-rate constant model was used to determine metabolic flux (K _FLT), blood to tissue transport (K ₁), and phosphorylation (k ₃). FDG-PET scans were obtained 75–90 min postinjection. Results   K _FLT and k ₃, but not K ₁ or k ₃/k ₂ + k ₃, reached significance for separating the recurrence from radionecrosis groups. Standardized uptake value and visual analyses of FLT or FDG images did not reach significance. Conclusions   K _FLT (flux) appears to distinguish recurrence from radionecrosis better than other parameters, FLT and FDG semiquantitative approaches, or visual analysis of images of either tracer.     

Effects of a 3‐day low‐fat diet on metabolic control,insulin sensitivity,lipids and adipocyte hormones in Norwegian subjects with hypertriacylglycerolaemia and type 2 diabetes

The metabolic and hormonal impact of rapid dietary changes in type 2 diabetes has not been clarified. The objective of this study was to test whether a short‐term, low‐fat diet affected metabolic control, insulin sensitivity, lipids and adipocyte hormones in patients with type 2 diabetes with hypertriacylglycerolaemia. Nineteen outpatient subjects (10 M, 9 F) with type 2 diabetes and triacylglycerols >2.2?mmol/L at screening were included in the study. Dietary intake was assessed by weighing during two periods of 3‐day baseline diet followed by a 3‐day low‐fat dietary intervention. The two periods of baseline diet did not differ with respect to relevant variables during intervention. Subjects were advised to increase fibre‐rich and low‐fat foods and to decrease intake of visible fat in an isoenergetic manner. The percentage of energy from fat was reduced from 39 to 22 (p<0.0001), median values. Daytime blood glucose did not change and fasting insulin and fasting glucose to insulin ratios were unaffected. Total cholesterol decreased from 6.3 to 6.2?mmol/L (p<0.005), high‐density lipoprotein cholesterol from 1.13 to 1.10?mmol/L (p<0.048) and the ratio of n‐6 to n‐3 fatty acids in phospholipids from 2.5 to 1.9 (p<0.003). Concentrations of leptin decreased from 12.1 to 9.9?ng/mL (p<0.005) and adiponectin increased from 8.6 to 10.5?μg/mL (p<0.024). The effect on leptin was confined to women. A low‐fat diet intervention for 3 days in insulin‐resistant type 2 diabetes affects lipid, adiponectin and leptin levels but fails to improve insulin sensitivity and metabolic control.     

Clinical viability of Fungitell,a new (1→3)-β-<Emphasis Type="SmallCaps">d</Emphasis>-glucan measurement kit,for diagnosis of invasive fungal infection,and comparison with other kits available in Japan

Fungitell, a (1→3)-β-d-glucan (β-d-glucan) measurement kit, was approved in the United States in 2004. Three other kits for measurement of β-d-glucan, Fungitec G test MK (G-MK), β-Glucan test Wako (Wako), and β-Glucan test Maruha (Maruha), are commonly used for diagnosis of invasive fungal diseases in Japan. We evaluated the clinical viability of the Fungitell kit and compared it with the 3 kits generally used in Japan. The plasma β-d-glucan values measured with each kit showed some differences, possibly because different β-d-glucan standards, blood pretreatment methods, and kinds of horseshoe crab (a raw material for the main reagent) are used in each kit. Measures of diagnostic efficiency, for example the sensitivity, specificity, and positive and negative predictive values, varied among the kits. Although the areas under the receiver operating characteristic curves of the kits were not significantly different, the sensitivity of the Fungitell kit was the highest, followed by that of the G-MK kit. The sensitivity of the Wako and Maruha kits was low, but the specificity of these tests was higher than that of the G-MK or Fungitell kits. These inconsistent β-d-glucan measurements could interfere with diagnosis of invasive fungal infection. Early establishment of an international standard method for measurement of β-d-glucan is required.