Innate lymphoid cells are reduced in pregnant HIV positive women and are associated with preterm birth.

Preterm birth is the leading cause of neonatal and child mortality worldwide. Globally, 1.4 million pregnant women are estimated to be living with HIV/AIDS, the majority of whom live in sub-Saharan Africa. Maternal HIV infection and antiretroviral treatment (ART) have been associated with increased rates of preterm birth, but the underlying mechanisms remain unknown. Acute HIV infection is associated with a rapid depletion of all three subsets of innate lymphoid cells (ILCs), ILC1s, ILC2s and ILC3s, which is not reversed by ART. ILCs have been found at the maternal-fetal interface and we therefore investigated the potential association between maternal HIV infection, peripheral ILC frequencies and preterm birth. In our study of pregnant South African women with accurately dated pregnancies, we show that maternal HIV infection is associated with reduced levels of all three ILC subsets. Preterm birth was also associated with lower levels of all three ILC subsets in early pregnancy. ILC frequencies were lowest in HIV positive women who experienced preterm birth. Moreover, ILC levels were reduced in pregnancies resulting in spontaneous onset of preterm labour and in extreme preterm birth (< 28 weeks gestation). Our findings suggest that reduced ILC frequencies may be a link between maternal HIV infection and preterm birth. In addition, ILC frequencies in early pregnancy may serve as predictive biomarkers for women who are at risk of delivering preterm.

γδ T cell frequencies are altered in HIV positive pregnant South African women and are associated with preterm birth.

BACKGROUND: Preterm birth is the leading cause of neonatal and child mortality worldwide. Maternal HIV infection and antiretroviral treatment (ART) increase the rate of preterm birth, but the underlying mechanisms remain unknown, limiting progress in prediction, prevention and treatment. While overall γδ T cell levels remain constant, acute HIV infection is associated with a depletion of the Vδ2 subset and an increase in the Vδ1 subset, which do not return to baseline with ART. γδ T cells have also been implicated in adverse pregnancy outcomes and we therefore investigated the potential association between maternal HIV infection, peripheral γδ T cell frequencies and preterm birth. METHODS: Study participants were HIV positive (n = 47) and HIV negative (n = 45) women enrolled in a prospective pregnancy cohort study at Chris Hani Baragwanath Academic Hospital in Soweto, South Africa. Women were enrolled in early pregnancy and gestational age was accurately determined by first trimester ultrasound scan. Peripheral blood samples were collected in each trimester and peripheral blood mononuclear cells isolated. Frequencies of γδ T cells, Vδ1+ and Vδ2+ γδ T cell subsets, and CCR6 chemokine receptor expression were determined by flow cytometry. RESULTS: Total γδ T cell levels were similar between HIV positive and HIV negative women throughout pregnancy. However, in each trimester maternal HIV infection was associated with reduced levels of the Vδ2+ subset and increased levels of the Vδ1+ subset, leading to a reversal of the Vδ1/Vδ2 ratio. Timing of ART initiation among HIV positive women did not affect levels of γδ T cells, the Vδ1+ and Vδ2+ subsets, or the Vδ1/Vδ2 ratio. Importantly, preterm birth was associated with lower total γδ T cell levels in early pregnancy and γδ T cell frequencies were lowest in HIV positive women who delivered preterm. Moreover, in the first trimester the proportion of Vδ1+ T cells that were CCR6+ was significantly reduced in HIV+ women and women who delivered preterm, resulting in the lowest proportion of CCR6+ Vδ1 T cells in HIV positive women who delivered preterm. CONCLUSIONS: Our findings suggest that altered γδ T cell frequencies may link maternal HIV infection and preterm birth. γδ T cell frequencies in early pregnancy may serve as predictive biomarkers to identify women at risk of delivering preterm.

Changes in the Vα7.2+ CD161++ MAIT cell compartment in early pregnancy are associated with preterm birth in HIV-positive women.

PROBLEM: Human immunodeficiency virus (HIV) infection is associated with an increased risk of adverse pregnancy outcomes, including preterm birth (PTB), despite viral suppression with antiretroviral therapy. Mucosal-associated invariant T (MAIT) cells are an immune cell subset involved in antimicrobial immunity at mucosal surfaces. MAIT cells have been found at the maternal-foetal interface, and MAIT cells are typically depleted early in HIV infection. We aimed to investigate changes in MAIT cells in relation to maternal HIV/ART status and PTB. METHOD OF STUDY: We conducted flow cytometric analysis of peripheral blood samples from 47 HIV-positive (HIV+) and 45 HIV-negative (HIV-) pregnant women enrolled in a prospective pregnancy cohort study in Soweto, South Africa. Frequencies of Vα7.2+ CD161++ MAIT cells and proportions of CD4+ , CD8+ and double-negative MAIT cells were compared between women with and without HIV infection, and between women with and without PTB or spontaneous preterm labour (Sp-PTL). RESULTS: Although overall MAIT cell frequencies were the same between HIV+ and HIV- patients, HIV+ patients had a higher proportion of CD8+ MAIT cells in the first two trimesters. Women with PTB and Sp-PTL also had a higher proportion of CD8+ MAIT cells in the first trimester compared to women without these outcomes. The association between changes in MAIT cell subsets and PTB/Sp-PTL was present in both HIV+ and HIV- women, and an additive effect on MAIT cell subsets was seen in women with both HIV infection and PTB. CONCLUSIONS: Interactions between HIV-related and pregnancy-related changes in MAIT cell subsets and distribution may lead to imbalances in peripheral MAIT cell subsets in early pregnancy. This may contribute to the increased risk of PTB in HIV+ patients by altering the overall functionality of the peripheral MAIT cell compartment.

Inhibitory Effects of Commonly Used Excipients on P-Glycoprotein in Vitro.

Pharmaceutical excipients are no longer considered inert and have been shown to influence the activity of metabolic enzymes and transporters, resulting in altered pharmacokinetics of substrate drugs. In this study, the effect of 25 excipients commonly used in drug formulations were investigated for their effect on P-glycoprotein (P-gp) activity. The effect of excipients on P-gp were assessed by measuring the change in the cellular accumulation of a P-gp substrate, digoxin, in MDCK-MDR1 (Madin Darby canine kidney transfected with multidrug resistance 1 gene) cells. The cells were exposed to low (10 µM) and high (200 µM) concentrations of excipient along with 10 µM digoxin. Excipient concentrations were chosen to span the range of concentrations previously used for investigating activities in vitro. At 10 µM of excipient, an increase in the intracellular digoxin concentration was seen with d-α-tocopherol poly-(ethylene glycol) succinate (Vit-E-PEG; p = 0.002), poly(ethylene oxide)20 sorbitan monooleate (Tween 80; p = 0.001), cetyltrimethylammonium bromide (CTAB; p = 0.021), poly(ethylene oxide)35 modified castor oil (Cremophor EL; p = 0.01), polyethylene glycol15-hydroxystearate (Solutol HS 15; p = 0.006), and poly(ethylene glycol) hexadecyl ether (Brij 58; p = 0.001). At 200 µM, Vit-E-PEG ( p < 0.0001), sodium 1,4-bis (2-ethylhexoxy)-1,4-dioxobutane-2-sulfonate (AOT; p < 0.0001), Tween 80 ( p < 0.0001), CTAB ( p = 0.004), poly(ethylene oxide)20 sorbitan monolaurate (Tween 20; p < 0.0001), Cremophor EL ( p < 0.0001), Solutol HS 15 ( p < 0.0001), Brij 58 ( p < 0.0001), and sodium carboxymethyl cellulose (NaCMC; p = 0.006) increased intracellular digoxin significantly. Concentration-dependent inhibition of P-gp was then investigated for selected excipients giving an IC50 for Vit-E-PEG (12.48 µM), AOT (192.5 µM), Tween 80 (45.29 µM), CTAB (96.67 µM), Tween 20 (74.15 µM), Cremophor EL (11.92 µM), Solutol HS 15 (179.8 µM), Brij 58 (25.22 µM), and NaCMC (46.69 µM). These data add to the growing body of evidence demonstrating that not all excipients are inert and will aid excipient choice for rational formulation development.

Derivation of CYP3A4 and CYP2B6 degradation rate constants in primary human hepatocytes: A siRNA-silencing-based approach.

The first-order degradation rate constant (kdeg) of cytochrome P450 (CYP) enzymes is a known source of uncertainty in the prediction of time-dependent drug-drug interactions (DDIs) in physiologically-based pharmacokinetic (PBPK) modelling. This study aimed to measure CYP kdeg using siRNA to suppress CYP expression in primary human hepatocytes followed by incubation over a time-course and tracking of protein expression and activity to observe degradation. The magnitude of gene knockdown was determined by qPCR and activity was measured by probe substrate metabolite formation and CYP2B6-Glo™ assay. Protein disappearance was determined by Western blotting. During a time-course of 96 and 60 h of incubation, over 60% and 76% mRNA knockdown was observed for CYP3A4 and CYP2B6, respectively. The kdeg of CYP3A4 and CYP2B6 protein was 0.0138 h-1 (±0.0023) and 0.0375 h-1 (±0.025), respectively. The kdeg derived from probe substrate metabolism activity was 0.0171 h-1 (±0.0025) for CYP3A4 and 0.0258 h-1 (±0.0093) for CYP2B6. The CYP3A4 kdeg values derived from protein disappearance and metabolic activity were in relatively good agreement with each other and similar to published values. This novel approach can now be used for other less well-characterised CYPs.

Bowel colonization with vancomycin-resistant enterococci after antimicrobial therapy for intra-abdominal infections: observations from 2 randomized comparative clinical trials of ertapenem therapy.

The impact of different antimicrobial regimens for intra-abdominal infections on the frequency of bowel colonization with vancomycin-resistant enterococci (VRE) was examined in 2 randomized open-label trials of intra-abdominal infection comparing piperacillin-tazobactam or ceftriaxone/metronidazole with ertapenem. In these short-term studies, overall rates of bowel colonization with VRE were generally comparable after treatment with piperacillin-tazobactam, ceftriaxone/metronidazole, or ertapenem.

Safety, tolerability, and immunogenicity of a two-dose regimen of high-titer varicella vaccine in subjects > or =13 years of age.

A new manufacturing process, known as process upgrade varicella vaccine (PUVV) was developed for a refrigerated formulation of varicella vaccine and for an investigational zoster vaccine. Safety and tolerability of a two-dose regimen of high-titered (approximately 50,000 PFU) PUVV were compared to a lower-titer formulation (approximately 5400 PFU) of VARIVAX; in 1366 healthy subjects > or =13 years old. Only one vaccine-related clinical serious adverse experience (pruritus; no hospitalization) was reported, in the VARIVAX group. Injection-site adverse experiences following any dose were higher in the PUVV group, 70.0%, than in the VARIVAX group, 56.2%, but generally were mild. Immunogenicity were similar in both groups in seronegative subjects. PUVV was generally well tolerated, and elicited an immune response similar to that induced by the marketed formulation of VARIVAX.

Efficacy and safety of ertapenem versus piperacillin-tazobactam for the treatment of intra-abdominal infections requiring surgical intervention.

Complicated intra-abdominal infections usually mandate prompt surgical intervention supplemented by appropriate antimicrobial therapy. The aim of this study was to demonstrate that ertapenem was not inferior to piperacillin-tazobactam for the treatment of community-acquired intra-abdominal infections. A randomized open-label active-comparator clinical trial was conducted at 48 medical centers on four continents from December 2001 to February 2003. Adult patients with intra-abdominal infections requiring surgery were randomized to receive either ertapenem 1 g daily or piperacillin/tazobactam 13.5 g daily in 3-4 divided doses. The primary analysis of efficacy was the clinical response rate in clinically and microbiologically evaluable patients at the test-of-cure assessment 2 weeks after completion of therapy. All treated patients were included in the safety analysis. Patient demographics, disease characteristics, and treatment duration in both treatment groups were generally similar. The most commonly isolated pathogens at baseline were E coli (greater than 50% of cases in each group) and B fragilis ( approximately 9%). Favorable clinical response rates were 107/119 (90%) for ertapenem recipients and 107/114 (94%) for piperacillin/tazobactam recipients. The frequencies of drug-related adverse events, most commonly diarrhea and elevated serum alanine aminotransferase levels, were similar in both treatment groups. Six of 180 ertapenem recipients (3%) and two of 190 piperacillin/tazobactam recipients (1%) had serious drug-related adverse experiences. In this study, ertapenem and piperacillin/tazobactam were comparably safe and effective treatments for adult patients with complicated intra-abdominal infections.

Ertapenem versus ceftriaxone and metronidazole as treatment for complicated intra-abdominal infections.

BACKGROUND: Prompt surgical intervention supplemented by appropriate antimicrobial therapy is usually required for successful treatment of complicated intra-abdominal infections. The objective of this study was to further evaluate the efficacy and safety of ertapenem relative to ceftriaxone/metronidazole as treatment for complicated intra-abdominal infections. METHODS: Adult patients with intra-abdominal infections requiring surgery were eligible for this open-label randomized trial comparing ertapenem 1 g daily with ceftriaxone 2 g daily plus metronidazole 30 mg/kg/day. The primary efficacy outcome was the clinical response rate in clinically and microbiologically evaluable participants at the test-of-cure (TOC) visit 2 weeks after discontinuation of therapy. All treated patients were included in the safety analysis. RESULTS: Participant demographics, disease characteristics, and duration of therapy in both treatment groups were generally similar. Escherichia coli was the most commonly isolated baseline pathogen, recovered in 52% of cases in each treatment group. Favorable clinical responses were achieved at TOC in 143 (96.6%) of 148 ertapenem recipients and in 146 (96.7%) of 151 ceftriaxone/metronidazole recipients. The frequencies of drug-related adverse events, most commonly nausea, diarrhea, vomiting, and elevated platelet count, were generally comparable in both treatment arms. Four ertapenem recipients (1.8%) and one ceftriaxone/metronidazole recipient (0.4%) experienced serious drug-related adverse events. CONCLUSIONS: In this study, ertapenem and ceftriaxone/metronidazole were comparably effective treatments for adult patients with complicated intra-abdominal infections.

Concomitant administration of a bivalent Haemophilus influenzae type b-hepatitis B vaccine, measles-mumps-rubella vaccine and varicella vaccine: safety, tolerability and immunogenicity.

BACKGROUND: The study was done to verify that concomitant administration of a bivalent Haemophilus influenzae type b-hepatitis B vaccine (Comvax), measles-mumps-rubella vaccine (M-M-RII) and varicella vaccine (Varivax) would be well-tolerated and suitably immunogenic with respect to all vaccine antigens. METHODS: We randomized 822 healthy 12- to 15-month-old children (1:1) to receive concomitant injections of Comvax, M-M-RII and Varivax (concomitant group) or Comvax followed 6 weeks later by injections of M-M-RII and Varivax (nonconcomitant group). Blood samples taken before and 6 weeks after vaccination were tested for antibodies to all vaccine antigens. RESULTS: Vaccinations were generally well-tolerated. Children in the concomitant and nonconcomitant treatment groups were similar with respect to the safety endpoint of primary interest (16.1 and 19.5%, respectively, had a fever > or =103 degree F rectally at any time within 14 days after either of two clinic visits). Fifteen serious adverse events were reported (eight in the concomitant group and seven in the nonconcomitant group); all resolved. Elements of two serious adverse events (fever, fever and measles-like rash; both in concomitant group children) were considered possibly related to vaccination. One child was withdrawn from the study because of a nonserious adverse event subsequently judged to be unrelated to vaccination. Similar proportions of vaccinees in the concomitant and nonconcomitant groups developed satisfactory antibody responses to the H. influenzae polysaccharide, polyribosylribitol phosphate (97.8 to 98.7%), hepatitis B surface antigen (99.2 to 100%), measles virus (99.4 to 99.6%), mumps virus (98.4 to 99.2%), rubella virus (100%) and varicella virus (93.2 to 94.6%). CONCLUSION: Concomitant administration of Comvax, M-M-RII and VARIVAX at the 12- or 15-month clinic visit is one satisfactory way of delivering some of the multiple vaccines indicated during the second year of life.

Decline in varicella-zoster virus (VZV)-specific cell-mediated immunity with increasing age and boosting with a high-dose VZV vaccine.

The safety and immunogenecity of a booster dose of live attenuated varicella-zoster virus (VZV) vaccine was evaluated in 196 healthy subjects, >or=60 years old, who had already received a VZV vaccine >5 years before. This repeat booster dose was well tolerated. Cell-mediated immunity (CMI) to VZV was measured by an interferon-gamma (IFN-gamma) enzyme-linked immunosorbent spot-forming cell (ELISPOT) assay and a limiting dilution responder cell frequency (RCF) assay. Prevaccination responses decreased as a function of increasing age but were detectable in all subjects by use of the IFN-gamma ELISPOT assay. In most subjects, VZV-specific CMI was increased at 6 weeks postvaccination. The magnitude of the vaccine-induced IFN-gamma ELISPOT response was inversely related to prevaccination values. Although there was a significant correlation between the IFN-gamma ELISPOT and RCF assays, the ELISPOT assay had greater sensitivity and a wider dynamic range. A live attenuated VZV vaccine is safe and immunogenic in an elderly population, and the vaccine-induced immunity may be monitored by the IFN-gamma ELISPOT assay.

Vaccination with measles, mumps and rubella vaccine and varicella vaccine: safety, tolerability, immunogenicity, persistence of antibody and duration of protection against varicella in healthy children.

BACKGROUND: Administration of M-M-R II (Measles, Mumps and Rubella Virus Vaccine, Live) and VARIVAX [Varicella Virus Vaccine Live (Oka/Merck)] given concomitantly at separate injection sites during the same office visit could increase vaccine compliance by reducing the number of health care visits for immunizations. We compared the safety and immunogenicity of M-M-R II and VARIVAX given concomitantly at separate sites (Group A) with administration of the two vaccines 6 weeks apart (Group B) as well as the persistence of varicella antibody and the duration of protection afforded by varicella vaccine. METHODS: A total of 603 healthy children, ages 12 months to 6 years, with no history of measles, mumps, rubella, varicella and zoster or vaccination against these diseases, were randomized to either Group A or B and were followed for clinical reactions and serologic responses to all four viral components. Children were enrolled from August through December, 1993. Subjects were followed for 5 years to evaluate persistence of varicella antibody and breakthrough varicella rates. We compared breakthrough rates to expected attack rates in unvaccinated children to produce estimates of vaccine efficacy. RESULTS: Both vaccine regimens were generally well-tolerated. There were no significant differences between the groups in the rates of fever, injection site reactions or rashes after vaccination. Seroconversion rates and geometric mean titers for measles, mumps and rubella were not significantly different between groups. The varicella seroconversion rate and percentage with glycoprotein-based ELISA titers > or = 5.0 units were similar between the two groups (99.5 and 92.5% vs. 100 and 94.8% for Groups A and B, respectively), but the geometric mean titers were statistically significantly different (13.2 for Group A and 17.9 for Group B). Varicella antibody persistence rates were >98 to 100% during 6 years of follow-up in both groups. Compared with historical rates, varicella vaccine efficacy during 5 years was estimated to be 90.5% (95% confidence interval, 86.2%, 95.0%) and 88.9% (95% confidence interval, 83.7%, 93.7%) in Groups A and B, respectively. CONCLUSIONS: Administration of M-M-R II and VARIVAX concomitantly at separate injection sites or 6 weeks apart was generally well-tolerated and immunogenic in healthy children 12 months to 6 years of age. VARIVAX administered with M-M-R II induced persistent immunity and long-term protection against breakthrough varicella infection.

A comparison of safety, tolerability and immunogenicity of Oka/Merck varicella vaccine and VARILRIX in healthy children.

This study compared safety, tolerability, and immunogenicity of the Oka/Merck varicella vaccine and VARILRIX [Oka-RIT strain SmithKline Beecham Biologicals] in healthy children 12-24 months of age. Subjects were randomized in this double blind study to receive either a single dose of Oka/Merck varicella vaccine, (approximately 50,000 plaque forming units (PFU), Group A or approximately 16,000 PFU, Group B) or 1 dose of VARILRIX, (approximately 40,000 PFU/dose, Group C). Safety profiles in each treatment group were similar. The proportions of subjects achieving a 6-week postvaccination titer> or = 5 gpELISA units in Groups A, B or C were 97.1, 95.2 and 85.6%, respectively.

Inverse relationship between six week postvaccination varicella antibody response to vaccine and likelihood of long term breakthrough infection.

BACKGROUND: We used the large clinical database that supported the development of Oka/ Merck varicella vaccine to study the relationship between the primary varicella antibody response, as determined by gpELISA, an enzyme-linked immunosorbent assay that detects antibodies to varicella-zoster virus (VZV) glycoprotein, and the subsequent risk of postvaccination breakthrough varicella. METHODS: We vaccinated 1,164 healthy children with a single dose of varicella vaccine containing 2900 to 9000 plaque-forming units/dose. The primary immune response to vaccination was determined by gpELISA 6 weeks after vaccination. Subjects were followed annually for 7 years to ascertain cases of breakthrough varicella. RESULTS: The estimated vaccine efficacy among children with a 6-week postvaccination antibody titer of > or = 5 gpELISA units was 95.5% (95% confidence interval, 94.2%, 96.8%) compared with 83.5% (95% confidence interval, 76.9%, 89.5%) for subjects with a titer of <5 gpELISA units. Children with a 6-week postvaccination antibody titer of <5 gpELISA units were 3.5 times more likely than those with a titer of > or = 5 gpELISA units to develop breakthrough varicella. CONCLUSIONS: We identified a 6-week postvaccination antibody titer of > or = 5 gpELISA units as an approximate correlate of protection. In addition we established an accelerated failure time model based on log normal hazard that predicted varicella breakthrough rates based on the distribution of 6-week postvaccination varicella antibody titers.