Ceftriaxone pharmacokinetics by a sensitive and simple LC-MS/MS method: Development and application.

Ceftriaxone is a third-generation cephalosporin, worldwide use as a first-line treatment for several infections, including life-threatening infections as meningitis or endocarditis. Nowadays, ceftriaxone use is changing, embracing high-dose schemes, new populations treated and requirement of dose individualization and optimization. These reasons warranted the development of new sensitive assays. This study aimed to develop and validate a fast and handy bioanalytical method for the quantification of ceftriaxone in human plasma covering a broad range of concentrations. The analysis was performed using high-performance liquid chromatography coupled to tandem mass spectrometry. Sample preparation was based on protein precipitation with acetonitrile followed by centrifugation. Chromatography separation was performed on Phenomenex Luna C18 column (5 µm, 150 × 2.0 mm) and a mobile phase consisting of 70 % of mobile phase A (10 mM of ammonium acetate and 1% formic acid in purified water) and 30 % mobile phase B (0.1 % formic acid in acetonitrile) at a flow rate of 500 µl/min on an isocratic program. Both the analyte and the internal standard were quantified using the positive electrospray ionization (ESI) mode within a single runtime of 5.00 min. The method was validated following the U.S. Food and Drug Administration guidelines over the concentration range of 3-1000 µg/mL. The within-run and between-run precision and accuracy were <15 %, and therefore met the standard regulatory acceptance criterion. In conclusion, a sensitive and robust LC-MS/MS method was developed for a fast quantitation of ceftriaxone concentrations in plasma samples with multiples applications in research and clinical therapeutic drug monitoring.

Ampicillin and Ceftriaxone Solution Stability at Different Temperatures in Outpatient Parenteral Antimicrobial Therapy.

The inclusion of ampicillin-containing regimens in outpatient parenteral antimicrobial therapy programs (OPAT) depends upon solution stability under conditions similar to those experienced in these programs. Lack of this information could hinder the inclusion in OPAT of patients suffering from Enterococcus faecalis infective endocarditis treated with ampicillin plus ceftriaxone. The purpose of this study is to determine the stability of ampicillin and ampicillin plus ceftriaxone solutions in a simulated outpatient setting conditions. Solutions of ampicillin 24 g/liter and ampicillin 24 g/liter combined with ceftriaxone 8 g/liter were stored at 25°C ± 2°C, 30°C ± 2°C and 37°C ± 2°C for 48 h. Chemical and physical stability were evaluated at 20, 24, 30, and 48 h after manufacturing. The solutions were considered stable if the percentage of intact drug was ≥90% and color and clearness remained unchanged. After 24 h of storage at a controlled temperature, ampicillin solution in 0.9% sodium chloride was found to be stable for 30 h at 25 and 30°C and for 24 h at 37°C. In the ampicillin plus ceftriaxone combined solution, both antibiotics were found to be stable after 30 h of storage at 25 and 30°C, but at 37°C, the stability criterion was not met at any time point. Our study offers solid evidence demonstrating that the concentrations of both drugs at two of the tested temperatures (25°C and 30°C) were stable for up to 30 h. Therefore, both ampicillin alone and ampicillin plus ceftriaxone solutions would be appropriate candidates for inclusion in OPAT programs.

Impact of universal access to hepatitis C therapy on HIV-infected patients: implementation of the Spanish national hepatitis C strategy.

In April 2015, the Spanish National Health System (SNHS) developed a national strategic plan for the diagnosis, treatment, and management of hepatitis C virus (HCV). Our aim was to analyze the impact of this on human immunodeficiency virus (HIV)-infected patients included in the HERACLES cohort during the first 6 months of its implementation. The HERACLES cohort (NCT02511496) was set up in March 2015 to evaluate the status and follow-up of chronic HCV infection in patients co-infected with HIV in the south of Spain. In September 2015, the data were analyzed to identify clinical events (death, liver decompensation, and liver fibrosis progression) and rate of treatment implementation in this population. The study population comprised a total of 3474 HIV/HCV co-infected patients. The distribution according to liver fibrosis stage was: 1152 F0-F1 (33.2 %); 513 F2 (14.4 %); 641 F3 (18.2 %); 761 F4 (21.9 %); and 407 whose liver fibrosis was not measured (12.3 %). During follow-up, 248 patients progressed by at least one fibrosis stage [7.1 %; 95 % confidence interval (CI): 6.3-8 %]. Among cirrhotic patients, 52 (6.8 %; 95 % CI: 5.2-8.9 %) developed hepatic decompensation. In the overall population, 50 patients died (1.4 %; 95 % CI: 1.1-1.9 %). Eight hundred and nineteen patients (23.56 %) initiated interferon (IFN)-free treatment during follow-up, of which 47.8 % were cirrhotic. In our study, during 6 months of follow-up, 23.56 % of HIV/HCV co-infected patients included in our cohort received HCV treatment. However, we observed a high incidence of negative short-term outcomes in our population.

Efficacy of Lysophosphatidylcholine in Combination with Antimicrobial Agents against Acinetobacter baumannii in Experimental Murine Peritoneal Sepsis and Pneumonia Models.

Immune response stimulation to prevent infection progression may be an adjuvant to antimicrobial treatment. Lysophosphatidylcholine (LPC) is an immunomodulator involved in immune cell recruitment and activation. In this study, we aimed to evaluate the efficacy of LPC in combination with colistin, tigecycline, or imipenem in experimental murine models of peritoneal sepsis and pneumonia. We used Acinetobacter baumannii strain Ab9, which is susceptible to colistin, tigecycline, and imipenem, and multidrug-resistant strain Ab186, which is susceptible to colistin and resistant to tigecycline and imipenem. Pharmacokinetic and pharmacodynamic parameters for colistin, tigecycline, and imipenem and the 100% minimal lethal dose (MLD100) were determined for both strains. The therapeutic efficacies of LPC, colistin (60 mg/kg of body weight/day), tigecycline (10 mg/kg/day), and imipenem (180 mg/kg/day), alone or in combination, were assessed against Ab9 and Ab186 at the MLD100 in murine peritoneal sepsis and pneumonia models. The levels of pro- and anti-inflammatory cytokines, i.e., tumor necrosis factor alpha (TNF-α) and interleukin-10 (IL-10), were determined by enzyme-linked immunosorbent assay (ELISA) for the same experimental models after inoculating mice with the MLD of both strains. LPC in combination with colistin, tigecycline, or imipenem markedly enhanced the bacterial clearance of Ab9 and Ab186 from the spleen and lungs and reduced bacteremia and mouse mortality rates (P < 0.05) compared with those for colistin, tigecycline, and imipenem monotherapies. Moreover, at 4 h post-bacterial infection, Ab9 induced higher TNF-α and lower IL-10 levels than those with Ab186 (4 µg/ml versus 3 µg/ml [P < 0.05] and 2 µg/ml versus 3.4 µg/ml [P < 0.05], respectively). LPC treatment combined with colistin, tigecycline, or imipenem modestly reduced the severity of infection by A. baumannii strains with different resistance phenotypes compared to LPC monotherapy in both experimental models.

Protease inhibitor monotherapy is effective in controlling human immunodeficiency virus 1 shedding in the male genital tract.

Cross-sectional study comparing seminal human immunodeficiency virus type 1 (HIV-1) shedding in patients receiving boosted protease inhibitor monotherapy (mtPI/rtv) (n = 66) versus triple therapy (TT) (n = 61). Seminal HIV-1 shedding rates in patients with undetectable plasma HIV-RNA were 16.0% on mtPI/rtv compared with 28.6% on TT (p 0.173). Aviraemic status and time on viral suppression were independently associated with lack of seminal HIV-1 shedding. During TT, non PI/rtv-based regimens were associated with a better control of HIV infection in semen despite similar time on viral suppression. The use of mtPI/rtv in well-controlled patients is not associated with increased seminal HIV excretion compared with TT.

Dimension of chronic hepatitis C virus in HIV-infected patients in the interferon-free era: an overview from south Spain.

The implementation of hepatitis C (HCV) direct-acting antiviral drugs is prioritized in several populations in which its application provides the most immediate and impactful benefit. In this scenario, a precise knowledge of the situation of human immunodeficiency virus (HIV)/HCV chronic co-infection is required to adequately address this disease. This cross-sectional study was performed in 21 hospitals in Andalusia (Spain). The study population consisted of HIV-infected patients with an active HCV chronic infection who were not receiving HCV treatment at the time of inclusion. A total of 13,506 HIV-infected patients were included in the study. Of them, 2561 (18.9 %) presented chronic HCV infection. The majority of the patients included were on highly active antiretroviral therapy (HAART; 96.2 %), showed plasma levels with an undetectable HIV viral load (92.5 %), and had a good immunological status (median CD4+ cell count of 486 cells/mL). The HCV genotype distribution was as follows: 58.1 % were genotype 1, 1.1 % were genotype 2, 16.1 % were genotype 3, and 22.1 % were genotype 4 (2.6 % were missing data). In total, 24.8 % of the patients showed liver fibrosis stage F0-F1, 27.9 % showed stage F2, 16.7 % showed stage F3, and 21 % showed stage F4 (9.6 % were missing data). With regards to previous HCV treatment experiences, 68.05 % of the patients were naïve and 31.95 % had failed to respond to a previous treatment. The burden of HCV/HIV co-infected patients in our population was reported as one in five HIV-infected patients requiring HCV treatment. The implementation of extra resources to face this important health challenge is mandatory.

Efficacy of and risk of bleeding during pegylated interferon plus ribavirin treatment in HIV/HCV-coinfected patients with pretreatment thrombocytopenia.

The aim of this study was to assess the efficacy of and the risk of major bleeding during pegylated interferon (peg-IFN)/ribavirin (RBV) treatment among human immunodeficiency virus (HIV)/hepatitis C virus (HCV)-coinfected patients according to the pretreatment platelet count. Two hundred and seventy-four HCV/HIV-coinfected, previously naïve individuals with compensated cirrhosis enrolled in one Spanish prospective cohort who received peg-IFN/RBV were included in this study. The frequency of severe bleeding and sustained virological response (SVR) rate were compared between patients with a pretreatment platelet count ≤70,000/mm(3) and >70,000/mm(3), respectively. Sixty-one (22 %) patients had a baseline platelet count ≤70,000/mm(3). The median (Q1-Q3) pretreatment platelet count was 58,000 (49,000-65,000) cells/mm(3) in the platelet ≤70,000 group and 129,000 (102,500-166,000) cells/mm(3) in the platelet >70,000 group (p < 0.0001). Seventeen (28 %) subjects of the platelet ≤70,000 group and 71 (33 %) patients of the platelet >70,000 group achieved SVR (p = 0.4). Only 2 (3.2 %) patients in the platelet ≤70,000 group developed a severe hemorrhagic event, specifically esophageal variceal bleeding. The efficacy of therapy with peg-IFN/RBV in HIV/HCV-coinfected patients with low pretreatment platelet counts is comparable to that found in the overall subset of subjects with compensated cirrhosis. The frequency of severe hemorrhagic events related with this therapy is low in this population.

Low concordance and resistance mutation emergence in the HIV protease gene among circulating and cell-associated viruses at viral replication episodes during darunavir/ritonavir monotherapy.

OBJECTIVE: To assess the changes on the HIV protease gene in plasma and peripheral blood mononuclear cell (PBMC) compartments during viral replication episodes in patients on boosted-darunavir monotherapy (mtDRV/rtv). METHODS: A prospective study was carried out in which adult HIV-1-infected patients who started mtDRV/rtv after viral suppression for ≥ 6 months with no major darunavir-related resistance mutations were enrolled. Patients with two consecutive plasma HIV RNA measurements >200 HIV-1 RNA copies/mL were considered as having virological failure (VF), while patients with two consecutive plasma HIV RNA measurements >50 copies/mL without meeting the VF criteria were considered to have virological rebound (VR). HIV protease genotypic profiles from plasma and PBMCs were performed at baseline and at VF and VR episodes. RESULTS: One hundred and fifty patients were included in the study, with overall VF and VR rates of 14% (n=21) and 14.7% (n=22), respectively. No major darunavir resistance mutations were observed in the plasma or PBMC samples. Circulating and cell-associated viruses showed a wild-type protease gene sequence in 54% and 23% of patients, respectively while the remainder patients only harboured minor protease inhibitor-associated mutations. Full concordance between plasma RNA and PBMC DNA protease genotypes was found in 23% of the sequences. CONCLUSIONS: No darunavir-related mutations were found in patients with VF or VR, either in plasma or in PBMCs; thus, simplification to mtDRV/rtv does not comprise future antiretroviral treatment options.

Impact of the peginterferon-α 2a and ribavirin plasma levels on viral kinetics and sustained virological response in genotype 1 HCV/HIV-co-infected patients with the unfavourable non-CC IL28B genotypes.

Studies on the association between the peginterferon-α and ribavirin levels and sustained virological response (SVR) have shown yielded conflicting results, but most of them were performed before the influence of IL28B polymorphisms was known. Our aim was to assess the effects of peginterferon-α 2a and ribavirin plasma levels on viral kinetics and SVR in hepatitis C virus genotype 1 HCV-1/HIV-co-infected patients according to IL28B genotype. This was a cohort study of HCV-1/HIV-co-infected patients who were HCV-treatment naïve and for whom the efficacy of peginterferon-α 2a plus ribavirin was evaluated by per-protocol analysis. The peginterferon-α 2a and ribavirin levels were measured by ELISA and HPLC-UV, respectively. The relationships among host and viral factors, the trough drugs levels and virological responses were analysed by multivariate regression analyses. A total of 131 Caucasian patients were included (cirrhosis:38.9%). Overall, SVR rate was 39.6%. In patients with CC IL28B genotype, SVR was related neither to peginterferon-α 2a nor to ribavirin plasma levels, while higher levels of both drugs were the only variables independently associated with SVR in individuals with CT/TT IL28B genotypes (OR, 5.02; CI95 , 1.45-17.1; P = 0.001 and 4.0; CI95 , 1.08-14.7; P = 0.038, respectively). Moreover, faster viral declines were observed in CT/TT patients when pegIFN-α 2a and ribavirin plasma levels were greater than 3400 pg/mL and 1.6 µg/mL, respectively. In contrast to the results for CC patients, the results in patients carrying the unfavourable CT/TT IL28B genotypes showed that plasma levels of both drugs have significant effects on viral kinetics and SVR.

Immune activation throughout a boosted darunavir monotherapy simplification strategy.

Our aim was to assess the evolution and the impact that blips, intermittent low-level viraemia and virological failure (VF) episodes have on patients' immune activation (IA) profiles during ritonavir-boosted darunavir monotherapy (mtDRV/rtv). A prospective cohort of human immunodeficiency virus-1-infected patients who switched to mtDRV/rtv was followed for 2 years. Cellular IA was assessed according to HLA-DR and CD38 expression in CD4(+) and CD8(+) T-cells and their naïve, effector memory and central memory subpopulations, and systemic IA was evaluated according to sCD14 and D-dimer levels. Seventy-five patients from the MonDAR cohort were selected for this substudy, and classified according to viral outcome as having continuous undetectable viraemia (n = 19), blips (n = 19), intermittent viraemia (n = 21), and VF (n = 16). The IA profile was closely linked to viral behaviour. Patients on viral suppression for 24 months showed a significant decrease in CD4(+) and CD8(+) T-cell activation and sCD14 and D-dimer levels. Patients with transient low-level viraemia episodes (blips and intermittent viraemia) showed cellular and systemic IA similar to baseline values. In contrast, significant increases in T-cell activation and sCD14 and D-dimer levels were observed in patients with VF. Baseline levels of HLA-DR(+)CD38(+)CD8(+) T-cells of >6.4% were independently associated with the emergence of VF. Therefore, mtDRV/rtv might be considered as a safe simplification strategy, on the basis of the IA results, whenever viral replication is under medium-term and long-term control. Transient low-level viraemia episodes do not affect patients' IA status. Moreover, HLA-DR(+)CD38(+)CD8(+) T-cell baseline levels should be considered when patients are switched to mtDRV/rtv.

Very low prevalence and no clinical significance of occult hepatitis B in a cohort of HIV-infected patients with isolated anti-HBc seropositivity: the BHOI study.

PURPOSE: Data on occult HBV infection in HIV patients are conflicting. We aimed to analyse the prevalence and clinical significance of occult hepatitis B in HIV-infected subjects. METHOD: An open-label, cross-sectional, multicentre study including all subjects with isolated anti-HBc seropositivity from a cohort of 3,030 HIV-infected patients was undertaken. HBsAg and HBsAb were both negative in all cases, and those patients with acute or convalescent hepatitis B were excluded. HBV DNA was quantified by PCR with a detection limit of 20 IU/mL. RESULTS: We found 5 cases (2.5%) of occult hepatitis B among 202 HIV-patients with isolated anti-HBc. The mean HBV DNA was 66 (15-112) IU/mL, none had symptomatic hepatitis, and their features, including aminotransferase levels, were similar to those without occult HBV infection. CONCLUSIONS: Occult hepatitis due to HBV is very unusual in HIV-positive patients with isolated anti-HBc. The use of standard regimens of HAART including drugs with activity against HBV might underestimate the prevalence of occult HBV infection. These patients had a very low viral load, no identifiable risk factors, and no greater risk of hypertransaminasaemia or the development of symptomatic hepatitis.

Role of pegylated interferon-alpha-2a and ribavirin concentrations in sustained viral response in HCV/HIV-coinfected patients.

The effect of simultaneous plasma concentrations of pegylated interferon-alpha-2a (pegIFN-alpha-2a) and ribavirin (Rbv) on viral response has not been addressed to date. Hepatitis C virus (HCV)/human immunodeficiency virus (HIV)-coinfected patients received pegIFN-alpha-2a and Rbv under routine clinical care conditions. Plasma concentrations of the two drugs were measured using enzyme-linked immunosorbent assay and high-performance liquid chromatography after 2, 4, 8, and 12 weeks and at the end of the treatment period (24-48 weeks, according to HCV genotype and treatment duration). Large interindividual variability was observed in the plasma levels of both drugs. After multivariate analysis, only HCV genotype 3, low HCV-RNA levels, and pegIFN-alpha-2a exposure remained as independent factors associated with sustained viral response (SVR). The probability of attaining an SVR in HCV genotypes 1 and 4 was more than three to four times higher in patients with pegIFN-alpha-2a levels above the selected cutoff point. Early therapeutic drug monitoring of pegIFN-alpha-2a levels could be beneficial in improving current treatment outcomes.

Influence of treatment and immunological recovery on tuberculosis relapses in HIV-infected patients.

SETTING: Studies on tuberculosis (TB) relapse in HIV-infected patients show contradictory results regarding the optimal duration of treatment. OBJECTIVE: To assess the incidence of TB relapse and associated factors in HIV-infected patients receiving a 9-month tuberculostatic regimen and concomitant HAART. PATIENTS AND METHODS: Observational prospective study recording 156 episodes of TB in 137 patients, most of whom were on a 9-month regimen of daily isoniazid and rifampicin-based TB treatment. The primary outcome measure was relapse after completion of therapy. RESULTS: Forty episodes were excluded due to death or loss to follow-up. The median follow-up was 24 months. Twenty-seven episodes of TB relapse were observed in 22 patients, yielding a relapse rate of 1.9/100 patient-years in those on a regimen of > or = 9 months. A high recurrence rate was observed in those who had prematurely suspended treatment. Treatment duration > or = 9 months and achieving both an undetectable viral load and increasing CD4-cell counts with HAART were associated with the absence of TB relapses. CONCLUSIONS: Considering its safety and tolerance, our results suggest that a 9-month regimen would be recommendable in patients with severe immunosuppression until the optimal duration of TB treatment in HIV-infected patients has been defined in a randomised clinical trial including HAART.

Simplicity and efficacy of a once-daily antiretroviral regimen with didanosine, lamivudine, and efavirenz in naïve patients: the VESD study.

PURPOSE: Our aim was to analyze the efficacy and safety of didanosine-lamivudine-efavirenz in a cohort of HIV patients starting antiretroviral therapy between January and September 2003. METHOD: We undertook a prospective, open-label, observational, multicenter study. RESULTS: 163 patients were enrolled. Over a 48-week period, plasma HIV RNA levels declined sharply, with a median decrease at the end of the observation time of >4.62 log copies/mL. The proportion of patients achieving a plasma HIV RNA level below 50 copies/mL was 62.9% (intention-to-treat analysis) at the end of the study period. The mean CD4 cell count increased steadily over time by 199 cells/microL. Antiviral efficacy was similar in patients with a baseline HIV RNA level above or below 100,000 copies/mL. Overall, 57 (34.1%) patients interrupted therapy; 9 due to lack of treatment response, 18 due to adverse side-effects, and 30 patients lost to follow-up or who withdrew their consent. Adherence was very high (90%-95%) and quality of life was good or very good in 69%. CONCLUSION: The once-daily combination of didanosine-lamivudine-efavirenz resulted in sustained viral suppression and was well-accepted by patients under real-life conditions, even immunosuppressed patients and those with a high viral load. Associated adverse events and virological failures were few.

Influence of the response to induction therapy on the rate of progression of cytomegalovirus retinitis in AIDS patients on intravitreal maintenance therapy.

In order to assess whether complete inactivation of retinitis at the end of induction therapy leads to delayed progression during maintenance therapy with weekly intravitreal ganciclovir, the time to the first progression to retinitis was evaluated in 27 AIDS patients (34 eyes) with stable cytomegalovirus retinitis. Data were censored before the introduction of protease inhibitors. Overall, retinitis progressed in 22 of 34 eyes in a median time of 12 weeks (mean +/- SD, 33 +/- 9 weeks). However, retinitis progressed in 15 of 19 eyes in which only partial inactivation was achieved following induction therapy (median time, 10 weeks; mean +/- SD, 17 +/- 4 weeks) but in only 7 of 15 eyes when complete inactivation was obtained (median time, 59 weeks; mean +/- SD, 56 +/- 19 weeks) (P= 0.02). There were no differences between the groups in CD4+ cell counts, drugs, route of induction treatment, or length of induction therapy. Induction therapy should be prolonged until complete inactivation of retinitis is obtained, since achieving only a partial response appears to be a factor in earlier progression when patients are switched to maintenance therapy with intravitreal ganciclovir.