Use of different inhibitory quotients to predict early virological response to tipranavir in antiretroviral-experienced human immunodeficiency virus-infected patients.

Information about the relationship between pharmacological parameters and an early virological response to tipranavir (TPV) is scarce. Human immunodeficiency virus (HIV)-infected patients who had received TPV as part of a salvage regimen were analyzed retrospectively. A virological response was defined as a decline in the HIV RNA level of > or = 1 log unit or to <50 copies/ml between weeks 4 and 12 of therapy. The virtual inhibitory quotient (vIQ) was calculated as the ratio of the TPV plasma trough concentration (C(trough))/virtual change in the 50% inhibitory concentration. Three genotypic inhibitory quotients (gIQs) were calculated by using different TPV resistance mutation scores (from the International AIDS Society-USA [IAS-USA], Randomized Evaluation of Strategic Intervention in Multidrug-Resistant Patients with Tipranavir [RESIST], and Agence Nationale de Recherches sur le Sida et les Hépatites Virales [ANRS] trials). The sensitivities, specificities, positive predictive values (PPVs), negative predictive values (NPVs), and likelihood ratios for a positive result (LHR+) and a negative result (LHR-) [LHR+ = sensitivity/(1-specificity); LHR- = (1-sensitivity)/specificity] were calculated. A total of 57 HIV-infected patients were analyzed. A virological response was achieved by 77% of the patients. TPV resistance mutations, TPV C(trough), vIQs, and gIQs were all significantly associated with a virological response. The vIQ had the best PPV and NPV (97% and 78%, respectively). The values of the LHR+ were 7.8 for vIQ, 3.4 for the RESIST gIQ, 3.3 for the IAS-USA gIQ, 3.1 for the ANRS gIQ, 2.2 for TPV C(trough), and 1.3 for the IAS-USA and RESIST scores. The values of LHR- were 0 for the RESIST score, 0.07 for vIQ, 0.09 for the IAS-USA score, 0.27 for the RESIST gIQ, 0.32 for the IAS-USA gIQ, 0.37 for the ANRS gIQ, and 0.48 for TPV C(trough). HIV-infected patients who initiate a salvage regimen based on TPV may benefit from baseline drug resistance testing and TPV plasma concentration determination, as vIQ is the best predictor of a virological response.

Correlation between human immunodeficiency virus type 1 (HIV-1) RNA measurements obtained with dried blood spots and those obtained with plasma by use of Nuclisens EasyQ HIV-1 and Abbott RealTime HIV load tests.

The plasma human immunodeficiency virus (HIV) RNA load is used in the clinical routine for the monitoring of HIV infection and the patient's response to antiretroviral therapy. Other body fluids or dried blood spots (DBS) can be used, however, to assess the level of viremia. The use of DBS may be especially helpful for the monitoring of HIV-infected patients in resource-poor settings, where access to adequate laboratory facilities is often difficult. However, the correlation between the HIV RNA levels in plasma and those in DBSs has not been well established. Paired plasma and DBS samples obtained from HIV type 1 (HIV-1)-infected patients were tested for HIV RNA copy numbers by using two different commercial assays, the Nuclisens EasyQ HIV-1 (version 1.1) test (the Nuclisens test; Biomerieux) and the m2000rt RealTime HIV test (the m2000rt test; Abbott). Nucleic acid extraction was performed manually by using either the Nuclisens isolation kit (which uses the Boom methodology) or the m2000rt sample preparation kit (an iron particle-based method). A total of 103 paired plasma and DBS samples were tested. Viral load results were obtained for 97 (94.2%) samples with the Nuclisens isolation kit and 81 (78.6%) samples with the m2000rt kit. The overall correlation between the RNA loads in plasma and DBS was good, although better results were obtained by the Nuclisens test (R(2) = 0.87, P < 0.001) than by the m2000rt test (R(2) = 0.70, P < 0.001). While the specificities were excellent and similar for both the Nuclisens and the m2000rt tests (97.1% and 100%, respectively), the sensitivity was greater by the Nuclisens test than by the m2000rt test (75.8% and 56.6%, respectively). Overall, the viral loads in DBS tended to be lower than those in plasma, with mean differences of 0.3 log unit (standard deviation, 0.5 log unit) and 0.76 log unit (standard deviation, 0.8 log unit) for the Nuclisens and the m2000rt tests, respectively. The levels of agreement between the measurements in plasma and DBS were assessed by using the Bland-Altman plot for each assay. The Nuclisens test gave results within its defined limits (-0.65 to 1.26) for 95.9% of the samples, while the m2000rt test gave results within its limits (-0.83 to 2.33) for 100% of the samples. In summary, the HIV-1 load can accurately be quantified by testing DBS by either the Nuclisens or the m2000rt test, although the Nuclisens test may outperform the m2000rt test when nucleic acids are extracted manually.

Changing patterns in HIV reverse transcriptase resistance mutations after availability of tenofovir.

Assessment of 1177 human immunodeficiency virus (HIV) resistance genotypes at an HIV/AIDS clinic showed a decrease in the incidence of the K65R mutation, from 15.2% of isolates during the period 2002-2004 to 2.7% of isolates during the period 2005-2006 (P < .001), despite elevated and stable rates of tenofovir use. A reduction in the rate of coadministration of didanosine (from 41.6% of patients in 2004 to 0.8% of patients in 2006; P < .001) largely explained this observation.

Preparation and Characterization of Antimicrobial Films Based on LDPE/Ag Nanoparticles with Potential Uses in Food and Health Industries.

In this work, the antimicrobial effect of silver nanoparticles in polyethylene based nanocomposites has been investigated using a non-conventional processing method to produce homogeneous materials. High energy ball milling under cryogenic conditions was used to achieve a powder of well-blended low-density polyethylene and commercial silver nanoparticles. The final composites in the form of films were obtained by hot pressing. The effect of various silver nanoparticles content (0, 0.5, 1 and 2 wt %) on the properties of low-density polyethylene and the antimicrobial effectiveness of the composite against DH5α Escherichia coli were studied. The presence of silver nanoparticles did not seem to affect the surface energy and thermal properties of the materials. Apart from the inhibition of bacterial growth, slight changes in the aspect ratio of the bacteria with the content of particles were observed, suggesting a direct relationship between the presence of silver nanoparticles and the proliferation of DH5α E. coli (Escherichia coli) cells. Results indicate that these materials may be used to commercially produce antimicrobial polymers with potential applications in the food and health industries.

Changing rates and patterns of drug resistance mutations in antiretroviral-experienced HIV-infected patients.

Surveillance of drug resistance mutations in antiretroviral-experienced HIV(+) patients may provide useful information regarding options available for rescue interventions. All resistance tests performed from 1999 to 2005 on antiretroviral-experienced individuals at one reference laboratory in Madrid were examined. Only mutations associated with drug resistance recorded at the September 2006 IAS-USA list were considered. A total of 2137 specimens were analyzed. Overall, 71.1% showed resistance mutations to at least one drug class, 56.1% to at least two, and 21% to all three drug families. Resistance mutations were 65% for NRTI, 44.4% for NNRTI, and 42.5% for PI. Mutations T215Y/F, M184V, and M41L were the most frequent for NRTI. Their rate significantly declined since 1999. K65R significantly increased since 1999 (0.8%) to 2003 (7.3%) but declined up to 3.3% in 2005. For NNRTI, K103N significantly increased from 21.8% in 1999 to 29.5% in 2005 (p < 0.01). The most frequent PI resistance mutations were L90M (24.3%), V82X (19.9%), M46I/L (19.5%), and I54V (17.1%). The presence of five or more was 58.8% in 1999 but declined to 22.2% in 2005. The rate of drug resistance mutations causing NRTI and PI resistance has steadily declined in antiretroviral-experienced patients since 1999. The availability of a large number and/or more convenient NRTI as well as the wide use of ritonavir-boosted PI could explain these observations. However, broad PI cross-resistance was seen in nearly 25% of antiretroviral-experienced patients in 2005. Therefore, there is a still need for new antiretrovirals with different resistance profiles.

Prevalence of the HIV-1 protease mutation I47A in clinical practice and association with lopinavir resistance.

Mutation proI47A has recently been associated with lopinavir/ritonavir (LPV/r) resistance. Only four out of 1859 specimens (0.2%) sent for drug resistance testing (219 drug-naive and 1650 antiretroviral-experienced) showed I47A. All belonged to patients failing LPV/r. The prevalence among protease inhibitor-experienced patients was 0.6%. Phenotypic testing showed that proI47A caused high-level lopinavir resistance (> 100-fold) and cross-resistance to amprenavir, whereas it caused hypersusceptibility to saquinavir. ProI47A should thus be considered the primary lopinavir resistance mutation.

Plasma levels of atazanavir and the risk of hyperbilirubinemia are predicted by the 3435C-->T polymorphism at the multidrug resistance gene 1.

The 3435C-->T polymorphism at the multidrug resistance gene 1 (MDR1) was examined in 74 patients with human immunodeficiency virus who initiated atazanavir therapy. The MDR1 genotype distribution at position 3435 was 28% CC, 45% CT, and 27% TT. Plasma levels of atazanavir were significantly higher in patients with genotype CC than in those with CT or TT, and bilirubin levels correlated with atazanavir concentrations.

Risk of selecting K65R in antiretroviral-naive HIV-infected individuals with chronic hepatitis B treated with adefovir.

Seven antiretroviral-naive HIV-infected individuals with chronic hepatitis B treated with adefovir for longer than 6 months were assessed. Using bulk population sequencing and a sensitive limiting dilution analysis, the selection of K65R or other resistance mutations did not occur in HIV, suggesting that adefovir can be confidently used as hepatitis B virus (HBV) therapy in HIV/HBV-co-infected patients who do not require antiretroviral therapy.

Antiretroviral recommendations may influence the rate of transmission of drug-resistant HIV type 1.

BACKGROUND: Human immunodeficiency virus (HIV) treatment guidelines have evolved, shifting from more-aggressive to more-conservative approaches. The potential impact of these shifts on the transmission of drug-resistant virus is unknown. METHODS: Drug-resistance genotypes were examined in all consecutive patients with recent HIV type 1 (HIV-1) seroconversion (hereafter, "HIV-1 seroconverters") seen at 10 Spanish hospitals since 1997. During the same period, the proportion of patients with chronic HIV-1 infection having undetectable viremia was examined, to estimate the extent and effectiveness of antiretroviral therapy. RESULTS: A total of 141 recent HIV-1 seroconverters were identified, 67.4% of whom were men who have sex with men. The rate of primary drug-resistance mutations, by year of infection, was 33.3% for 1997, 29.4% for 1998, 20% for 1999, 14.3% for 2000, 3.4% for 2001, 15.4% for 2002, and 10.9% for 2003. On the other hand, the proportion of 8388 persons with chronic HIV-1 carriage who had an undetectable virus load was 33.4% for 1997, 34.6% for 1998, 39.7% for 1999, 47.5% for 2000, 52.9% for 2001, 39.7% for 2002, and 58.1% for 2003. A significant inverse correlation between transmission of drug-resistant HIV-1 and undetectable virus load was found (r=-0.955, by Spearman's test; P=.001). The lowest rate of transmission of drug-resistant HIV-1 was seen in 2001, when relatively "aggressive" treatment guidelines were used. Transmission of drug-resistant HIV-1 increased in 2002, in parallel with a reduction in the number of patients with chronic HIV-1 carriage and undetectable virus load, reflecting the popularity of drug holidays or treatment interruptions. CONCLUSION: The rate of drug resistance in recent HIV-1 seroconverters inversely correlates with the proportion of chronically HIV-1-infected individuals who have undetectable virus loads in the same region, which indirectly reflects antiretroviral treatment rules at any given time.

Resistance to nonnucleoside reverse-transcriptase inhibitors and prevalence of HIV type 1 non-B subtypes are increasing among persons with recent infection in Spain.

The prevalence of drug resistance mutations was 12.1% among 198 persons who experienced human immunodeficiency virus (HIV) seroconversion identified in Spain during 1997-2004. There was a significant increase of K103N and of non-B subtypes over time. Transmission of HIV infection around the time of seroconversion was shown in 8 couples and in 2 clusters of 3 individuals.

Higher efavirenz concentrations determine the response to viruses carrying non-nucleoside reverse transcriptase resistance mutations.

We examined the influence of both efavirenz plasma concentrations and non-nucleoside reverse transcriptase (NNRTI) resistance mutations on the antiviral activity of efavirenz in patients experiencing early virological failure under nevirapine-containing regimens. Up to 41% of patients reach less than 50 copies/ml at 48 weeks. No association was found between the presence of NNRTI resistance mutations and virological outcome. Nevertheless, patients responding virologically and carrying NNRTI-resistant viruses had higher efavirenz levels than those who did not respond.

Predictors of selection of K65R: tenofovir use and lack of thymidine analogue mutations.

Over the past 5 years, 1846 HIV-infected patients underwent drug resistance testing at our institution. None out of 216 drug-naive subjects showed K65R. However, it was recognized in 53 out of 1630 antiretroviral-experienced patients (3.3%), of whom 10 had never been exposed to tenofovir. The rate of K65R increased from 0.6% in 1999 to 11.5% in 2004. The recognition of K65R correlated negatively with the presence of thymidine analogue mutations but positively with Q151M.

Decline in the rate of genotypic resistance to antiretroviral drugs in recent HIV seroconverters in Madrid.

Genotypic resistance to antiretroviral drugs was analysed in plasma from 57 acute or recent HIV seroconverters in Madrid. The overall prevalence of drug-associated primary resistance mutations was 25.8% in 1997-1999, but declined to 3.8% in 2000-2001. The lower rate in recent years suggested that most new HIV infections derive from viraemic individuals unaware of their HIV- positive status, rather than from those failing antiretroviral treatment. Drug-resistance testing is thus not required before beginning antiretroviral therapy.

Evidence for differences in the sexual transmission efficiency of HIV strains with distinct drug resistance genotypes.

BACKGROUND: The transmission of drug-resistant human immunodeficiency virus type 1 (HIV-1) is a matter of major concern, because it could compromise the response to antiretroviral therapy. MATERIAL AND METHODS: From 1997 through 2003, genotypic drug resistance profiles in 89 patients with recent HIV-1 seroconversion were compared in a case-control study with HIV-1 genotypes obtained from 520 subjects identified at Hospital Carlos III in Madrid, Spain, as "potential transmitters" of drug-resistant virus. This group consisted of HIV-infected patients experiencing virologic failure of antiretroviral therapy. RESULTS: Drug resistance mutations were recognized in 15 (16.8%) of 89 patients with recent HIV-1 seroconversion (the seroconverter group), providing resistance to nucleoside reverse-transcriptase inhibitors (NRTIs) in 14.6%, nonnucleoside reverse-transcriptase inhibitors (NNRTIs) in 3.4%, and protease inhibitors (PIs) in 3.4%. Among individuals who were potential transmitters of HIV-1 (the potential transmitter group), drug resistance mutations were found in 80%, providing resistance to NRTIs in 73.7%, NNRTIs in 36.4%, and PIs in 38.7%. The estimated ratio of individuals with recent HIV-1 seroconversion to potential transmitters for drug-resistance genotypes was 0.23 for resistance to NRTIs, 0.09 for resistance to NNRTIs, and 0.09 for resistance to PIs. For specific resistance mutations, the ratio of individuals with recent HIV-1 seroconversion to potential transmitters was 0.18 for 41L, 0.20 for 215Y/F (including revertants), 0.06 for 184V, 0.04 for 103N, and 0.14 for 181C when considering drug resistance mutations in the reverse transcriptase (RT) gene; the ratio was 0.12 for 46L, 0.06 for 82A/T/S, and 0.08 for 90L, when examining PI resistance mutations. CONCLUSION: Strains of HIV with some drug resistance genotypes (41L, 215Y/F, and 181C in the RT gene and 46L in the protease gene) may be more efficiently transmitted than strains with other drug resistance mutations (184V and 103N in the RT gene and 82A/S/T and 90M in the protease gene).

Prevalence of G333D/E in naive and pretreated HIV-infected patients.

A substitution at codon 333 (G -->D/E) within the reverse transcriptase (RT) gene causes resistance to both zidovudine (AZT) and lamivudine (3TC) in a background of mutations associated with loss of sensitivity to both drugs. In the absence of G333D/E, M184V restores the sensitivity to AZT in viruses harboring AZT-resistant genotypes. In this study, we examined the prevalence of the G333D/E mutation in plasma samples from 401 HIV-infected individuals from a routine clinical practice. Genotypic results could be obtained in 277 (81%) specimens belonging to pretreated subjects and in all 65 samples from treatment-naive individuals. The overall prevalence of the G333D/E mutation was 13.2%, without significant differences when comparing naive (10.8%) and treatment-experienced patients (13.7%). The codon 333 mutation was associated with AZT/3TC-resistant genotypes in 76.3% of pretreated individuals but in none of the treatment-naive individuals. In conclusion, the G333D/E mutation must be considered a natural polymorphism, which appears in 11% of treatment-naive HIV-infected individuals. In AZT-experienced patients, it might be advisable to exclude its presence before adding 3TC in combination in the hope of obtaining a restoration of AZT sensitivity if M184V develops.

Indinavir plasma concentrations and resistance mutations in patients experiencing early virological failure.

Virological failure under protease inhibitor (PI)-based antiretroviral regimens is often not explained by the selection of resistance mutations. The role of low indinavir (IDV) plasma levels in treatment failure was assessed in 46 subjects experiencing early virological failure to a first-line IDV-containing triple combination. Overall, 69% of patients showed subtherapeutic IDV plasma levels (it was not detected at all in 75% of them). Subjects with detectable but suboptimal IDV levels developed more IDV resistance mutations. Thus, drug monitoring may be useful to assess treatment adherence and risk of drug resistance in early virological failures. This information may be crucial for choosing the most appropriate rescue intervention.

Correlation between rules-based interpretation and virtual phenotype interpretation of HIV-1 genotypes for predicting drug resistance in HIV-infected individuals.

Drug resistance testing provides useful information for managing HIV-infected patients. Phenotyping could add complementary information to genotyping and occasionally be more useful, although is less available to clinicians. Large paired geno-pheno databases have allowed the prediction of phenotypes from genotypes. However, the accuracy of these virtual phenotypes (vPT) in a clinical setting has not been well assessed yet. We analyzed the concordance between vPT and interpreted genotype (GT) in 105 samples belonging to treatment-experienced HIV-infected patients. A high concordance was seen when examining both non-nucleoside reverse transcriptase inhibitors (NNRTI) and protease inhibitors (PI) (r = 0.95 either), while it was lower for nucleoside analogs (r = 0.79). The drugs with lower concordance were abacavir (71.1%), tenofovir (71.5%) and didanosine (71.9%). In 20% of specimens (21/105), the vPT did not provide results for all approved drugs. These were mainly samples with a high number of drug resistance mutations or rare genotypes, which seem to be underepresented in the VircoNET database. Overall, there is good correlation between vPT/GT, especially for PI and NNRTI. The inclusion of additional sequences in the VircoNET database, mainly those derived from heavily treatment-experienced patients and/or from patients failing the most recently approved drugs might improve its performance.

Short communication high risk of endothelial dysfunction in HIV individuals may result from deregulation of circulating endothelial cells and endothelial progenitor cells.

Endothelial progenitor cells (EPC) and circulating endothelial cells (CEC) have recently been considered as biomarkers of cardiovascular risk (CVDR) in healthy subjects. The impact of HIV infection on these cells is not well known. A case-control study was conducted in 15 antiretroviral-naive HIV(+) patients and 15 HIV-negative controls. The quantitative profile of CEC and EPC differed significantly in HIV(+) and HIV(-) subjects. HIV(+) subjects had significantly more CEC and less EPC than HIV(-) controls. A quantitative impairment in the balance of the CEC and EPC might contribute to the increased subclinical CVDR in HIV(+) patients.

Minority HIV mutation detection in dried blood spots indicates high specimen integrity and reveals hidden archived drug resistance.

BACKGROUND: Dried blood spots (DBS) could serve as an attractive, cost-effective alternative to plasma for HIV drug resistance testing. OBJECTIVES: To assess the utility and potential gain in genotypic information with sensitive testing of DBS compared to conventional bulk plasma genotyping, and examine the correlation of majority and minority-level resistance mutations in DBS with treatment history. STUDY DESIGN: Evaluate nucleic acids from the DBS of 33 antiretroviral-experienced subtype B-infected subjects for minority M41L, K65R, K70R, K103N, Y181C, M184V, and T215Y/F mutations by real-time PCR. Compare minority resistance mutations in DBS with bulk genotypes from the same DBS cards and available plasma specimens. RESULTS: All but one (50/51, 98%) mutation from the original plasma bulk sequencing were still detectable in the DBS after three years of storage. The one mutation not identified in DBS was also no longer detectable by bulk sequencing. Furthermore, sensitive testing found 12 additional drug resistance mutations at minority levels in the DBS of 11 (33%) patients. Six minority mutations were in the RNA compartment and six were detected only in the DNA compartment. Resistance was detected in the DBS RNA compartment only in cases where the associated drug was in use within one year of sample collection. CONCLUSIONS: Our ability to identify majority and additional minority-level resistance mutations demonstrated that DBS, if stored properly, is a high-integrity specimen type for conventional and sensitive drug resistance testing. Our data further support the global utility of DBS for drug resistance surveillance and clinical monitoring.