170,025 research outputs found
Characterization of HIV-1 pol sequences in newly HIV-1 diagnosed patients: identification of drug resistance markers and their involvement in the transmission events
No full textAd oggi sono disponibili 25 farmaci per il trattamento dell’infezione da HIV. L’uso combinato di tali farmaci, la cosiddetta Highly Active Antiretroviral Therapy (HAART), ha consentito di migliorare le prospettive e la qualità di vita dei pazienti, rendendo possibile la soppressione della replicazione virale nella maggior parte dei soggetti infetti.
Tuttavia, se la terapia non riesce a sopprimere adeguatamente il virus, si osserva l’insorgenza di ceppi virali farmaco-resistenti in grado di compromettere inesorabilmente l’efficacia dei farmaci stessi. Attualmente, la presenza di virus farmaco-resistenti è messa in evidenza attraverso il test di resistenza genotipica che prevede il sequenziamento dei geni bersaglio dei farmaci antiretrovirali. Recentemente, sono state inoltre messe a punto delle metodiche in grado di rilevare ceppi resistenti anche in forma minoritaria, presenti cioè con una prevalenza inferiore al 20% nell’intera popolazione virale, il cui utilizzo nella pratica clinica è però ancora compromesso dai costi elevati. La più nota tra tutte è sicuramente il pyrosequencing.
Un problema strettamente connesso all’insorgenza di ceppi resistenti è rappresentato dalla loro trasmissione (sia come specie predominante che in forma minoritaria) a soggetti HIV-infetti che non hanno mai assunto alcun farmaco antiretrovirale (definiti drug-naïve). Attualmente, nei paesi occidentali, la prevalenza di virus resistente nei pazienti drug-naïve è stimata intorno al 10%, e diversi studi hanno evidenziato come tali ceppi (anche quando presenti in specie minoritarie) possano compromettere il successo della prima linea terapeutica.
In base a quanto riportato, l’obiettivo di questa tesi è stato quindi quello di caratterizzare la distribuzione e la diffusione della resistenza trasmessa in Italia, con particolare attenzione alle dinamiche di trasmissione della stessa e al ruolo che i soggetti drug-naïve rivestono in questo fenomeno.
Un ulteriore obiettivo è stato quello di definire nuovi marcatori genetici di HIV rilevabili attraverso metodiche standard di sequenziamento in grado di predire la presenza di varianti minoritarie resistenti nella popolazione virale. L’individuazione di mutazioni “sentinella” in grado di predire la presenza di quasispecie minoritarie resistenti riveste un ruolo centrale nel miglioramento dell’interpretazione genotipica e nella selezione di pazienti potenzialmente portatori di farmaco-resistenza.
Analizzando 255 pazienti di nuova diagnosi e naïve alla terapia antiretrovirale, abbiamo riscontrato che il 5,9% di questi mostrava resistenza trasmessa; in particolare il 3,9% era resistente alla classe farmacologica degli analoghi nucleosidici della trascrittasi inversa (NRTI), il 3,5% alla classe dei non-NRTI e lo 0,4% alla classe degli inibitori della proteasi (PI). Il 3,5% dei pazienti era inoltre portatore di un virus resistente a più classi farmacologiche. Comparando pazienti con infezione recente a pazienti con infezione cronica, abbiamo riscontrato che la prevalenza di virus farmaco-resistente era leggermente più bassa nella prima categoria rispetto alla seconda (3,4% versus 6,6%, anche se accompagnata da un P=0,09 non significativo), riflettendo probabilmente l’uso negli ultimi anni di farmaci sempre più potenti ed ad alta barriera genetica. Infine, la via di trasmissione che più delle altre sembrava incidere nella trasmissione della farmaco-resistenza era l’omosessualità (OR 7,7; 95% CI: 1,7–35,0, P=0,008).
Nella seconda parte di questa tesi, mediante metodiche filogenetiche avanzate, abbiamo caratterizzato i clusters epidemiologici di infezione in una coorte di 884 pazienti HIV-positivi, di cui 306 naïve alla terapia e 578 trattati. Da questa analisi è emerso come benché la fonte principale di trasmissione di virus farmaco-resistente fosse rappresentata dai pazienti in fallimento terapeutico, ben il 38,5% dei virus resistenti figurava in clusters composti esclusivamente da individui drug-naïve con infezione sia recente che cronica, sostenendo il ruolo di tali pazienti quale ulteriore fonte di farmaco-resistenza.
Nella terza parte di questa tesi abbiamo identificato alcune mutazioni nella trascrittasi inversa di HIV, la cui presenza nel test di resistenza genotipico correlava in modo significativo con specie minoritarie farmaco resistenti. In particolare, l’analisi di di 40 pazienti drug-naïve ha mostrato come il 70% di pazienti con la mutazioni L210M presentasse specie minoritarie farmaco-resistenti (P=0.03), supportando il ruolo di tale mutazione come “sentinella” di farmaco-resistenza nascosta.
In conclusione, questa tesi ha consentito di caratterizzare la resistenza trasmessa e le dinamiche di trasmissione di ceppi HIV-farmacoresistenti in Italia. Queste informazioni sono di fondamentale importanza per impostare programmi di sorveglianza e prevenzione dell’infezione da HIV. Tale tesi ha portato inoltre all’identificazione di mutazioni “sentinella” in grado di predire la presenza di specie minoritarie resistenti non rilevabili dai test genotipici utilizzati comunemente nella pratica clinica. Ciò è di fondamentale importanza per ottimizzare la gestione clinica del pazienti infetto da HIV.To date 25 drugs are available for the HIV-1 treatment. The combined use of these drugs, known as Highly Active Antiretroviral Therapy (HAART) has successfully suppressed the HIV-1 replication and has dramatically improved the prognosis of HIV-1 infected patients.
However, if viral rebound occurs during therapy, viruses with mutations conferring drug resistance can be selected. To date, drug resistant viruses can be detected by the genotypic resistance test, based on the sequencing of the genes target of the antiretroviral therapy. Recently, new assays (as ultra deep pyrosequencing) have allowed to assess antiretroviral drug resistance even when present as minority species (with a prevalence <20% of the entire viral population). However, their use is still limited in clinical practice due to their high cost.
The frequently selection of drug resistant strains in treatment failing patients can in turn increase the risk of their transmission (both in predominant and both in minority species) to HIV-1 infected individuals, naïve to the antiretroviral drugs (drug-naïve). To date, in countries with a wide access to HAART, the prevalence of drug resistant strains in drug-naïve patients is around 10%, and many studies have demonstrated that these drug resistance strains (also in minority species) are associated with an increased probability of virological failure to the first-line antiretroviral therapy.
In this light, the aim of this thesis was to characterize the distribution and the spread of HIV-1 drug resistance in Italy, with particular attention to the population dynamics of transmitted resistance and the role of untreated patients in the spread of drug-resistance. The last objective of this thesis was to define new genetic markers of HIV that can predict the presence of transmitted drug resistant minority species. The identification of these “sentinel” mutations could improve the genotypic interpretation and could help the selection of patients potentially reservoir of drug-resistance.
Among 255 newly diagnosed and untreated individuals, the 5.9% of patients showed signs of transmitted resistance; in particular, 3.9% of patients was infected with nucleoside reverse-transcriptase inhibitors (NRTI)-resistant viruses, 3.5% with non-NRTI-resistant viruses and 0.4% with protease inhibitors-resistant viruses. In addition, the 3.5% of patients carried HIV-1 resistant strains with more than one major drug resistance mutation. Comparing chronic infections with recent infections, we also found a decreased rate of resistance in recent infections (P=0.09), reflecting the increasing use of potent drugs and highly active antiretroviral regimens in recent years. Homosexual individuals were also more likely to harbour a virus with at least one primary resistance mutation (OR 7.7; 95% CI: 1.7–35.0, P=0.008).
In the second part of this thesis, we have investigated the epidemiological networks characterizing the HIV-1 infection by using a phylogenetic approach in a cohort of 884 patients. Of them, 306 were drug-naïve and 578 were HAART-treated individuals. Even if patients failing HAART remained the principal source of transmitted drug resistance, the 38.5% of drug resistant viruses was involved in clusters composed only by drug-naïve individuals, supporting the role of these patients as drug-resistance transmitters.
In the third part of this thesis, we have characterized specific mutations in the reverse-transcriptase of HIV, those presence significantly correlated with minority drug resistance variants. In particular, among the 40 drug naïve patients analyzed, we found that the 70.0% of patients with L210M carried drug resistance minority species (P=0.03), supporting the role of this mutation as “sentinel” of hidden drug resistance mutations.
In conclusion, this thesis was able to characterize the HIV-1 transmitted drug resistance and the transmission dynamics of HIV-1 drug resistant strains in Italy. These data were essential to improve the surveillance and the preventive programmes on HIV infections. This thesis has also characterized “sentinel” mutations able to predict minority drug resistance variants, undetectable by standard genotypic tests. Taking into account these genetic markers right from diagnosis can help clinicians to improve the management of HIV-1 infected patients
Recent Decisions
No full textComments on recent decisions by Harry Contos, Jr., A. Alteri, William A. Loy, Richard D. Schiller, Karl Jorda, and Richard C. Clark
Recent Decisions
No full textComments on recent decisions by Harry Contos, Jr., A. Alteri, William A. Loy, Richard D. Schiller, Karl Jorda, and Richard C. Clark
Recent Decisions
No full textComments on recent decisions by Harry Contos, Jr., A. Alteri, William A. Loy, Richard D. Schiller, Karl Jorda, and Richard C. Clark
Sequencing Analysis in the Field of HIV Variability and Drug Resistance
No full textSequence analysis plays a crucial role in routine clinical practice, especially in the field of HIV-1 drug resistance. Resistance testing in HIV-1 infected patients is now recommended to guide the choice of antiretroviral therapy in clinical practice. In this regard, genotypic testing is the most common method used for detecting drug-resistant strains of HIV. It measures indirectly the drug resistance by detecting mutations in HIV genome known to be associated with a reduced susceptibility to antiviral drugs either in the laboratory and/or in clinical situations (http://www.iasusa.org/resistance_mutations; http://hivdb.stanford.edu). Various tools are now available to assist the provider in interpreting genotypic test results. Many clinicians currently use on a routine basis the genetic information on HIV-genotype to make treatment decisions for HIV-infected patients (both drug-naive or drug-experienced). Sequences obtained by genotypic assays can also be used to evaluate the HIV-1 genetic variability. Indeed, it should be considered that antiretroviral drug design, resistance research, and interpretation systems have been largely based on HIV-1 subtype B, which has historically been the most prevalent in North America, Western Europe, and Australia. However, subtype B viruses account for only about 12% of the worldwide HIV-1 infections. An increasing number of individuals who are infected with non-B subtype strains are now receiving antiretroviral therapy because of rollout programs in resource-limited world and because of increasing migration to the developed world, particularly to European countries. Therefore, HIV-1 subtype determination is essential to appreciate the effects of HIV-1 genetics on the characteristics of drug susceptibility and drug resistance. Finally, genotypic assays can be used for surveillance purposes both in developed and resource-constrained countries. Regarding these last settings, a careful evaluation of HIV drug resistance is needed since the number of antiretroviral drug-experienced patients is rapidly increasing. In light of previous considerations, in the present chapter book we reported the results obtained in two already published studies (Fokam et al., 2011; Ceccarelli et al., 2012). The HIV-1 genetic diversity and the prevalence of resistance among Cameroonian individuals were assessed trough the analysis of HIV-1 sequences. Sequences from 239 adults and 92 children (including both drug-naive and drug-experienced patients) were performed and analyzed. In particular, drug resistance mutations in HIV-1 pol region (corresponding to the entire protease and the first 280/300 amino acids of the reverse transcription open reading frame) were characterized; phylogenetic analysis was performed for subtype determination. Among 40 children failing first-line antiretroviral therapy, treatment response was also evaluated at weeks 24 and 48 after treatment, on the basis of genotypic results. The phylogenetic analysis revealed a great genetic diversity of HIV-1 viral strains circulating in Cameroon. The totality of the analyzed population was infected by non-B subtypes. In particular, CRF02_AG was the most common viral subtype (about 60% both in children and adults). Among drug-naive patients drug resistance was rather low (8.2% in adults and 4.9% in children), while a very high prevalence of resistance was observed in patients failing therapy (about 80% in adults and 90% in children). The high level of drug resistance observed in the analyzed cohort of treated patients is alarming, since it occurred as a result of few years of treatment. Adherence to therapy, adequate physicians’ education and the appropriate use of genotypic resistance testing are the critical points of intervention to improve patient’s care. For example, in the analysis performed in children starting a second-line regimen after failure, treatment change driven by genotypic resistance test was successful for most of them, highlighting the importance of choosing a therapy according to the genetic characteristics of the virus. Overall, these data well describe the role of HIV-1 genotypic resistance test in clinical practice. This testing has revolutionized the care of HIV infected patients and significantly advanced HIV research. Making use of this assay, physicians can closely monitor the emergence of drug resistance mutations and optimize the management of patients infected with drug resistant HIV. To date, important efforts are addressed to the improvement of these tests, in term of sensitivity, cost-effectiveness and detection of drug resistance in non-B subtypes. The introduction of new techniques for next generation sequencing (detecting also minority species representing 1% of viral population) and of new affordable algorithms to evaluate the importance of resistance patterns for non-subtype B strains is crucial to achieve this goal
The Role of HIV Infection in Neurologic Injury
No full textThe central nervous system (CNS) is a very challenging HIV-1 sanctuary, in which HIV-1 replication is established early on during acute infection and can persist despite potent antiretroviral treatments. HIV-1 infected macrophages play a pivotal role acting as vehicles for HIV-1 to spread into the brain, and can be the major contributor of an early compartmentalization. HIV-1 infection in CNS may lead to a broad spectrum of neurological syndromes, such as dementia, mild neurocognitive disorders, and asymptomatic impairment. These clinical manifestations are caused by the release of neurotoxins from infected cells (mainly macrophages), and also by several HIV-1 proteins, able to activate cell-signaling involved in the control of cellular survival and apoptosis. This review is aimed at highlighting the virological aspects associated with the onset of neurocognitive disorders and at addressing the novel therapeutic approaches to stop HIV-1 replication in this critical sanctuary
P1896 Relationship between virological evolution and acute exacerbation of chronic hepatitis C (A–E–CHC) and during the natural history of infection
No full textRelationship between virological evolution and acute exacerbation of chronic hepatitis C (A-E-CHC) and during the natural history of infection
No full textGoing Beyond Counting First Authors in Author Co-citation Analysis
No full textThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Mitomycin C in highly myopic eyes - Author reply
No full textOphthalmology. 2005 Feb;112(2):208-18; discussion 219.
Mitomycin C modulation of corneal wound healing after photorefractive keratectomy in highly myopic eyes.
Gambato C, Ghirlando A, Moretto E, Busato F, Midena E.
SourceRefractive Surgery Service and Antimetabolite Therapy Research Unit, Department of Ophthalmology, University of Padova, Padova, Italy.
Abstract
PURPOSE: To evaluate the role of topical mitomycin C in corneal wound healing (CWH) after photorefractive keratectomy (PRK) in highly myopic eyes.
DESIGN: Prospective, double-masked, randomized clinical trial.
PARTICIPANTS: Seventy-two eyes of 36 patients affected by high (>7 diopters) myopia.
METHODS: In each patient, one eye was randomly assigned to PRK with intraoperative topical 0.02% mitomycin C application, and the fellow eye was treated with a placebo. Postoperatively, mitomycin C-treated eyes received artificial tears (3 times daily, tapered in 3 months), whereas the fellow eye was treated with fluorometholone sodium 2% and artificial tears (3 times daily, tapered in 3 months).
MAIN OUTCOME MEASURES: Uncorrected visual acuity (UCVA) and best-corrected visual acuity (BCVA), contrast sensitivity, manifest refraction, and biomicroscopy. Contrast sensitivity was determined using the Pelli-Robson chart. Corneal confocal microscopy documented CWH.
RESULTS: Mean follow-up was 18 months (range, 12-36). No side effects or toxic effects were documented. At 12-month follow-up examination, UCVAs (logarithm of the minimum angle of resolution) were 0.4+/-0.48 and 0.5+/-0.53 (P = .03) in mitomycin C-treated eyes and corticosteroid-treated eyes, respectively. At 1 year, corneal haze developed in 20% of corticosteroid-treated eyes, versus 0% of mitomycin C-treated eyes. At 12, 24, and 36 months, corneal confocal microscopy showed activated keratocytes and extracellular matrix significantly more evident in untreated eyes (Ps = 0.004, 0.024, and 0.046, respectively).
CONCLUSION: Topical intraoperative application of 0.02% mitomycin C can reduce haze formation in highly myopic eyes undergoing PRK.
Comment in
Ophthalmology. 2006 Feb;113(2):357; author reply 357-8
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