Methodological needs in the quality and safety characterisation of nanotechnology-based health products: Priorities for method development and standardisation.

Nanotechnology-based health products are providing innovative solutions in health technologies and the pharmaceutical field, responding to unmet clinical needs. However, suitable standardised methods need to be available for quality and safety assessments of these innovative products prior to their translation into the clinic and for monitoring their performance when manufacturing processes are changed. The question arises which technological solutions are currently available within the scientific community to support the requested characterisation of nanotechnology-based products, and which methodological developments should be prioritized to support product developers in their regulatory assessment. To this end, the work presented here explored the state-of-the-art methods to identify methodological gaps associated with the preclinical characterisation of nanotechnology-based medicinal products and medical devices. The regulatory information needs, as expressed by regulatory authorities, were extracted from the guidance documents released so far for nanotechnology-based health products and mapped against available methods, thus allowing an analysis of methodological gaps and needs. In the first step, only standardised methods were considered, leading to the identification of methodological needs in five areas of characterisation, including: (i) surface properties, (ii) drug loading and release, (iii) kinetic properties in complex biological media, (iv) ADME (absorption, distribution, metabolism and excretion) parameters and (v) interaction with blood and the immune system. In the second step, a detailed gap analysis included analytical approaches in earlier stages of development, and standardised test methods from outside of the nanotechnology field that could address the identified areas of gaps. Based on this analysis, three categories of methodological needs were identified, including (i) method optimisation/adaptation to nanotechnological platforms, (ii) method validation/standardisation and (iii) method development for those areas where no technological solutions currently exist. The results of the analysis presented in this work should raise awareness within the scientific community on existing and emerging methodological needs, setting priorities for the development and standardisation of relevant analytical and toxicological methods allowing the development of a robust testing strategy for nanotechnology-based health products.

Modelling the long-acting administration of anti-tuberculosis agents using PBPK: a proof of concept study.

SETTING: Anti-tuberculosis formulations necessitate uninterrupted treatment to cure tuberculosis (TB), but are characterised by suboptimal adherence, which jeopardises therapeutic efficacy. Long-acting injectable (LAI) formulations or implants could address these associated issues. OBJECTIVE: niazid, rifapentine, bedaquiline and delamanid-in adults for treatment for latent tuberculous infection (LTBI). DESIGN: PBPK models were developed and qualified against available clinical data by integrating drug physicochemical properties and in vitro and population pharmacokinetic data into a mechanistic description of drug distribution. Combinations of optimal dose and release rates were simulated such that plasma concentrations were maintained over the epidemiological cut-off or minimum inhibitory concentration for the dosing interval. RESULTS: The PBPK model identified 1500 mg of delamanid and 250 mg of rifapentine as sufficient doses for monthly intramuscular administration, if a formulation or device can deliver the required release kinetics of 0.001-0.0025 h-1 and 0.0015-0.0025 h-1, respectively. Bedaquiline and isoniazid would require weekly to biweekly intramuscular dosing. CONCLUSION: We identified the theoretical doses and release rates of LAI anti-tuberculosis formulations. Such a strategy could ease the problem of suboptimal adherence provided the associated technological complexities for LTBI treatment are addressed.

Long-acting formulations for the treatment of latent tuberculous infection: opportunities and challenges.

Long-acting/extended-release drug formulations have proved very successful in diverse areas of medicine, including contraception, psychiatry and, most recently, human immunodeficiency virus (HIV) disease. Though challenging, application of this technology to anti-tuberculosis treatment could have substantial impact. The duration of treatment required for all forms of tuberculosis (TB) put existing regimens at risk of failure because of early discontinuations and treatment loss to follow-up. Long-acting injections, for example, administered every month, could improve patient adherence and treatment outcomes. We review the state of the science for potential long-acting formulations of existing tuberculosis drugs, and propose a target product profile for new formulations to treat latent tuberculous infection (LTBI). The physicochemical properties of some anti-tuberculosis drugs make them unsuitable for long-acting formulation, but there are promising candidates that have been identified through modeling and simulation, as well as other novel agents and formulations in preclinical testing. An efficacious long-acting treatment for LTBI, particularly for those co-infected with HIV, and if coupled with a biomarker to target those at highest risk for disease progression, would be an important tool to accelerate progress towards TB elimination.

The Effect of Gene Variants on Levonorgestrel Pharmacokinetics When Combined With Antiretroviral Therapy Containing Efavirenz or Nevirapine.

Reduced levonorgestrel concentrations from the levonorgestrel contraceptive implant was previously seen when given concomitantly with efavirenz. We sought to assess whether single nucleotide polymorphisms (SNPs) in genes involved in efavirenz and nevirapine metabolism were linked to these changes in levonorgestrel concentration. SNPs in CYP2B6, CYP2A6, NR1I2, and NR1I3 were analyzed. Associations of participant demographics and genotype with levonorgestrel pharmacokinetics were evaluated in HIV-positive women using the levonorgestrel implant plus efavirenz- or nevirapine-based antiretroviral therapy (ART), in comparison to ART-naïve women using multivariate linear regression. Efavirenz group: CYP2B6 516G>T was associated with lower levonorgestrel log10 Cmax and log10 AUC. CYP2B6 15582C>T was associated with lower log10 AUC. Nevirapine group: CYP2B6 516G>T was associated with higher log10 Cmax and lower log10 Cmin . Pharmacogenetic variations influenced subdermal levonorgestrel pharmacokinetics in HIV-positive women, indicating that the magnitude of the interaction with non-nucleoside reverse transcriptase inhibitors (NNRTIs) is influenced by host genetics.

Prevalence and type of drug-drug interactions involving ART in patients attending a specialist HIV outpatient clinic in Kampala, Uganda.

OBJECTIVES: Scale-up of HIV services in sub-Saharan Africa has rapidly increased, necessitating evaluation of medication safety in these settings. Drug-drug interactions (DDIs) involving antiretrovirals (ARVs) in sub-Saharan Africa are poorly characterized. We evaluated the prevalence and type of ARV DDIs in Ugandan outpatients and identified the patients most at risk. METHODS: A total of 2000 consecutive patients receiving ARVs at the Infectious Diseases Institute, Kampala were studied. The most recent prescription for each patient was screened for clinically significant DDIs using www.hiv-druginteractions.org. Univariable and multivariable logistic regression were used to identify risk factors for DDIs. A screening tool was developed using significant risk factors and tested in a further 500 patients. RESULTS: Clinically significant DDIs were observed in 374 (18.7%) patients, with a total of 514 DDIs observed. Only 0.2% of DDIs involved a contraindicated combination. Comedications commonly associated with DDIs were antibiotics (4.8% of 2000 patients), anthelmintics (2.2%) and antifungals (3.5%). Patient age, gender, CD4 count and weight did not affect risk of DDIs. In multivariable analysis, the patient factors that independently increased risk of DDIs were two or more comedications (P < 0.0001), a PI-containing ARV regimen (P < 0.0001), use of an anti-infective (P < 0.0001) and WHO clinical stage 3-4 (P = 0.04). A scoring system based on having at least two of these risk factors identified between 75% and 90% of DDIs in a validation cohort. CONCLUSIONS: Significant ARV DDIs occur at similar rates in resource-limited settings and developed countries; however, the comedications frequently causing DDIs differ. Development of tools that are relevant to particular settings should be a priority to assist with prevention and management of DDIs.

Successful pharmacogenetics-based optimization of unboosted atazanavir plasma exposure in HIV-positive patients: a randomized, controlled, pilot study (the REYAGEN study).

BACKGROUND: Atazanavir without ritonavir, despite efficacy and tolerability, shows low plasma concentrations that warrant optimization. METHODS: In a randomized, controlled, pilot trial, stable HIV-positive patients on atazanavir/ritonavir (with tenofovir/emtricitabine) were switched to atazanavir. In the standard-dose arm, atazanavir was administered as 400 mg once daily, while according to patients' genetics (PXR, ABCB1 and SLCO1B1), in the pharmacogenetic arm: patients with unfavourable genotypes received 200 mg of atazanavir twice daily. EudraCT number: 2009-014216-35. RESULTS: Eighty patients were enrolled with balanced baseline characteristics. The average atazanavir exposure was 253 ng/mL (150-542) in the pharmacogenetic arm versus 111 ng/mL (64-190) in the standard-dose arm (P < 0.001); 28 patients in the pharmacogenetic arm (75.7%) had atazanavir exposure >150 ng/mL versus 14 patients (38.9%) in the standard-dose arm (P = 0.001). Immunovirological and laboratory parameters had a favourable outcome throughout the study with non-significant differences between study arms. CONCLUSIONS: Atazanavir plasma exposure is higher when the schedule is chosen according to the patient's genetic profile.

Pharmacokinetic and pharmacodynamic analysis of efavirenz dose reduction using an in vitro-in vivo extrapolation model.

The pharmacokinetics (PK) of efavirenz (EFV) is characterized by marked interpatient variability that correlates with its pharmacodynamics (PD). In vitro-in vivo extrapolation (IVIVE) is a "bottom-up" approach that combines drug data with system information to predict PK and PD. The aim of this study was to simulate EFV PK and PD after dose reductions. At the standard dose, the simulated probability was 80% for viral suppression and 28% for central nervous system (CNS) toxicity. After a dose reduction to 400 mg, the probabilities of viral suppression were reduced to 69, 75, and 82%, and those of CNS toxicity were 21, 24, and 29% for the 516 GG, 516 GT, and 516 TT genotypes, respectively. With reduction of the dose to 200 mg, the probabilities of viral suppression decreased to 54, 62, and 72% and those of CNS toxicity decreased to 13, 18, and 20% for the 516 GG, 516 GT, and 516 TT genotypes, respectively. These findings indicate how dose reductions might be applied in patients with favorable genetic characteristics.

The use of trough ribavirin concentration to predict sustained virological response and haematological toxicity in HIV/HCV-co-infected patients treated with ribavirin and pegylated interferon.

OBJECTIVES: To study the association between trough ribavirin concentration (C(trough)) with sustained virological response (SVR) and haemoglobin (Hb) decrease in HIV/hepatitis C virus (HCV)-co-infected (HIV+/HCV+) patients treated with anti-HCV therapy. METHODS: HIV+/HCV+ patients treated with ribavirin and pegylated interferon were prospectively evaluated. Qualitative and quantitative HCV-RNA, Hb levels and ribavirin C(trough) were measured at baseline and weeks 2, 4, 12, 24, 36 and 48 during therapy. HCV-RNA was also measured at 24 weeks after the end of therapy. Efficacy analysis was performed on patients with a definitive virological outcome (SVR, relapser and non-responder), whereas for toxicity analysis, dropouts were considered until the last available observation. RESULTS: Fifty-two patients (54.7% with genotype 1 or 4) were included. Overall, no correlation between ribavirin C(trough) and early virological response (EVR) nor SVR was found. However, in patients with genotype 1 or 4, ribavirin C(trough) was independently associated with EVR (P = 0.036) and SVR (P = 0.046). A ribavirin C(trough) cut-off of 1600 ng/mL was found to be associated with both EVR (chi(2) = 5.69, P = 0.028) and SVR (chi(2)=4.2, P = 0.04). Higher ribavirin C(trough) correlated with Hb decrease (R = -0.361, P = 0.009) and was independently associated with an Hb decrease of >4 g/dL (P = 0.009). Receiver operating characteristic (ROC) analysis indicated that a ribavirin C(trough) of >2300 ng/mL was associated with an Hb decrease of >4 g/dL (chi(2) = 8.08, P = 0.01). CONCLUSIONS: Our study confirmed a relationship between ribavirin exposure and both efficacy and toxicity. Moreover, we found ribavirin C(trough) cut-offs for both SVR in genotypes 1 and 4 and overall haematological toxicity. These findings deserve further clinical evaluation.

Alpha-interferon as induction and maintenance therapy in hairy cell leukemia: a long-term follow-up analysis.

Although in recent years the use of purine analogues has increased the percentage of long-term complete response the effect on overall survival of patients with hairy cell leukemia (HCL) is not yet clear. This study aimed to evaluate the long-term outcome (mean follow up of 92 months) of 64 patients receiving IFN as first-line therapy. IFN was well tolerated and effective. The overall response rate was 91% (PR 65%, CR 13%, GPR 13%). Forty-one patients (63%) received IFN 3 MU/ wk as maintenance therapy. The 10-yr projected survival rate of responding patients (CR and GPR 100%; PR 95%) and non-responders (SD, PD 80%) clearly shows that type of response does not affect survival. Patients receiving IFN maintenance had a statistically higher PFS than those who did not (p <0.01). This study shows that IFN is still one of the standard therapies for this disease, that achieving CR has no primary relevance for the control of the disease, and that good utilization of therapeutic resources may assure HCL patients a survival rate comparable to that of a normal, healthy population.

Virtues and problems of the high C-field Cs beam frequency standard.

The high C-field Cs beam frequency standard is presently a working machine that is undergoing first evaluations. The projected 10(-14 ) accuracy goal is as yet unattained, mainly because of inadequate C-field uniformity and stability. An analysis of the projected possible C-field improvements and the consequent uncertainty is here reported.

First-trimester screening for Down syndrome using nuchal translucency measurement with free beta-hCG and PAPP-A between 10 and 13 weeks of pregnancy--the combined test.

In a population of 1467 women attending the 'G. Gaslini' Institute for antenatal care, we evaluated first-trimester risk screening for Down syndrome using the 'combined test' based on ultrasound measurement of nuchal translucency (NT), maternal serum pregnancy-associated plasma protein A (PAPP-A) and free beta-hCG, and maternal age. No clinical action was taken on these results. The gestational age, determined by scan measurement of crown rump length, ranged from 10 weeks to 13 weeks 6 days. The median maternal age was 31 years 8 months. There were 13 Down syndrome pregnancies. The risk of having an affected pregnancy was estimated from a multivariate Gaussian distribution, using commercially available software. With a risk cut-off of 1 in 350, 11 affected pregnancies were detected (detection rate 85 per cent, 95 per cent confidence interval: 56-100 per cent) with a 3.3 per cent false-positive rate. The odds of being affected given a positive result were 1 in 30. Further data are needed to determine, with greater statistical reliability, the relative performance of the combined test with current second-trimester screening.

Full-blown hypothyroidism associated with vitiligo and acropachy. Report of one case.

During the first two years after the onset of hypothyroidism, a patient with spontaneous primary myxedema developed thyroid acropachy. Fifteen years before the diagnosis of thyroid disease, he had patchy vitiligo of the face. There were high serum levels of antibodies against thyroglobulin (anti-Tg) and microsomal antigen (anti-M) were present, while the serum levels of antibodies against the second antigen of the colloid and the cell-surface antigen fell within the normal range. Moreover, antibodies to gastric parietal cells, adrenocortical cells or pancreatic islets in the serum were not present. Concerning the immuno-genetic pattern of our patient, his HLA system did not appear to confirm the well documented prevalence in whites with autoimmune disorders of an antigen specificity positive for the locus DR3.

Gastric histomorphology and secretory biochemistry after biliopancreatic diversion.

When paired data are considered and unpaired results are compared with controls, it appears that no significant changes of gastric histomorphology occur following biliopancreatic diversion.

Long-term hyposensitization and adverse immunologic responses. A laboratory evaluation.

Ninety atopic patients with allergic asthma and/or rhinitis were classified into three groups: group A, consisting of 30 patients never subjected to immunotherapy; group B, 30 patients receiving immunotherapy for various allergies for 3 to 24 months; group C, 30 patients receiving immunotherapy for 5 or more years. The following studies were performed using standard techniques: quantitative determination of serum immunoglobulin concentrations IgA, IgG, IgM, rheumatoid factor, C3 and C4 concentrations, antinuclear antibodies (ANA). Circulating immune complexes were detected by a competitive immuno-enzyme assay. No statistical differences were found in the groups studied for every parameter considered. Although signs and symptoms of autoimmune disease were not present, an elevated incidence of ANA in the entire group (23.3%) was found.

[Autoantibodies in advanced old-age]. / Autoanticorpi nell'eta' avanzata.

A variety of autoantibodies (anti-nuclear, anti-mitochondrial, anti-smooth muscle, anti-gastric parietal cell) were measured by indirect immunofluorescence. At least one of such autoantibodies was detected in 65.2% of 46 old people sera (ranging from 76 to 95). Both a decrease in the efficiency of the immune response and a failure of the immunoregulatory mechanisms occurring with age may account for the above findings.