Current HIV Research (v.9, #3)

Application of SCR Priming VLP Boosting as a Novel Vaccination Strategy Against HIV-1 by Seyed Mehdi Sadat, Rezvan Zabihollahi, Mohammad Reza Aghasadeghi, Rouhollah Vahabpour, Seyed Davar Siadat, Arash Memarnejadian, Kayhan Azadmanesh, Kazem Parivar (140-147).
Human immunodeficiency virus infection is a worldwide health problem and a protective vaccine is desperatelyneeded to control the AIDS pandemics. To address this concern, we previously constructed single-cycle replicable(SCR) HIV-1 virions, which completely maintained the antigenic structures of HIV-1. Herein, to optimize a vaccinationstrategy, we studied the immunogenicity of produced SCR virions and adjuvant-formulated HIV-1 virus-like particles(VLPs) in homologous and heterologous prime-boosting regimens. Accordingly, BALB/c mice received three doses ofimmunogens in 3-week intervals and their immune responses were evaluated using ELISA, cytokine and IFN-.. ELISpotassays. These analyses not only indicated the superiority of SCR prime-VLP boosting for strong induction of specificIFN-.. producing cells, but also showed the capability of this strategy over the others for better stimulation of humoralresponse, which was evidenced with the detection of highest titer of total IgG against HIV ENV glycoprotein.Furthermore, determination of IgG subclasses and IFN-../IL4 secretion ratio in cultured splenocytes demonstrated theefficient augmentation of mixed responses with the dominancy of Th1 immunity following SCR/VLP immunizationstrategy. Our results additionally pointed towards the applicability of Montanide ISA 720 + CpG as a potent Th1-directingadjuvant mixture. Overall, this study suggests SCR prime-VLP boosting as a promising approach in HIV vaccinedevelopment.

HIV Persistence in the Gut Mucosa of HIV-Infected Subjects Undergoing Antiretroviral Therapy Correlates with Immune Activation and Increased Levels of LPS by Gabriella d'Ettorre, Mirko Paiardini, Lorenzo Zaffiri, Mauro Andreotti, Giancarlo Ceccarelli, Cecilia Rizza, Marileda Indinnimeo, Stefano Vella, Claudio M. Mastroianni, Guido Silvestri, Vincenzo Vullo (148-153).
We investigated the relationship between viral persistence in the gut, microbial translocation, and T cellactivation during chronic HIV infection. Plasma levels of LPS, fraction of circulating CD8+CD38+ T cells, and levels ofHIV-DNA in rectosigmoid biopsies and peripheral blood mononuclear cells were determined in 22 HIV-infectedindividuals and 10 healthy controls. We found that in untreated HIV-infected individuals, HIV-DNA load was higher inthe gut mucosa than in the blood. Also, ART-treated patients exhibited lower levels of LPS and CD8+CD38+ T cells thanuntreated patients, but higher levels than controls. In ART-treated individuals, the level of HIV-DNA in the gut correlatedwith levels of LPS and fraction of CD8+CD38+ T cells. We concluded that in ART-treated individuals, higher levels ofgut-associated HIV-DNA are associated with persistent immune activation and microbial translocation.

Drug Resistance Mutations During Structured Interruptions of HAART in HIV-1-Infected Children by Gerardo C. Palacios-Saucedo, Lydia G. Rivera-Morales, Jose M. Vazquez-Guillen, Luz M. Sanchez, Teresa J. Ramirez, Evangelina Briones, Rocio Ortiz-Lopez, Carlos A. Vazquez, Cristina Rodriguez-Padilla (154-159).
Information concerning structured treatment interruptions (STI) of the Highly Active Antiretroviral Therapy(HAART) and the associated risk of this strategy for selecting antiretroviral resistance in children is scarce. In this study,we have searched for antiretroviral resistance mutations at the end of five viral rebounds of two HIV-1-infected childrenwho underwent an STI program. The HAART of two children with HIV and a chronically undetectable viral load (VL)was interrupted for 4 weeks and then restarted and continued for 12 weeks for three cycles. VL, CD4+/CD8+lymphocytes, and clinical status were evaluated at the end of each STI and at 6 and 12 weeks after HAART was resumed.Treatment of both the patients based on AZT+3TC+RTV remained identical during the study. The reverse transcriptase(RT)- and protease (PR)-coding regions were sequenced searching for antiretroviral resistance mutations at the end ofeach viral rebound. Genotyping analysis was performed using the Stanford University HIV Drug Resistance Database(HIV-DB). One patient experienced progressively lower viral rebounds (269000-31300 at the first and third rebounds,respectively), while the other patient did not experience such a reduction, and the VL of both the patients fell toundetectable levels during therapy. In the five viral rebounds examined, no mutations for major or minor resistance toprotease inhibitors (PIs) were found and the analysis indicated susceptibility to all PIs currently in clinical use. Althoughthe mutation K103R associated with non-nucleoside reverse transcriptase inhibitor (NNRTI) resistance was found in twoviral rebounds of one patient, the analysis indicated the absence of resistance to reverse transcriptase inhibitors in all thesequences evaluated. As no mutation related to antiretroviral resistance was found, our results suggest that the STIprogram used in this study may have a low risk of selecting antiretroviral resistance. Nevertheless, further studiesevaluating larger cohorts over longer periods, and above all, controlled clinical trials, are required before definitiveconclusions about the safety of STI of HAART in children may be drawn.

Prospective Observation for Seven-Year's Highly Active Antiretroviral Therapy in Chinese HIV-1 Infected Patients by Mei He, Yu-Huang Zheng, Hua-Ying Zhou, Diallo Mamadou, Zi Chen, Xia Chen, Yun-Hai Yao, Yan He (160-165).
To prospectively observe the efficacy, tolerability, immune reconstitution and toxicity of long-term highlyactive antiretroviral therapy (HAART) in Chinese patients infected HIV. 437 cases originally received two nucleosidereverse transcriptase inhibitors (NRTIs) and one non-nucleoside reverse transcriptase inhibitor (NNRTI) during a meanperiod of 4.3 years (3.1-7.3). Patients were followed up by HIV RNA levels, T lymphocyte subsets, blood routine test, andbiochemical parameters. If active opportunistic infections, apparent side effects or virological failure appeared,appropriate treatment would be taken immediately. 30 patients (6.86%) died, most in the first 6 months of HAART. Theproportion of subjects with HIV-1 RNA <500 copies/ml was 90.8%, 63.5%, 69.4%, 70.0% and 72.2% at 1, 4, 5, 6 and 7year. The CD4+ T cell count was 115, 246, 301, 334, 363, 356,386 and 373 cells/ul at 0, 1, 2, 3, 4, 5, 6 and 7 year. 67.9%showed various drug-related side effects, most including gastrointestinal side-effects, nervous disorder, myelotoxicity andabnormal liver function, rashes, serum cholesterol elevation, mostly appearing in the first 12 months. Grade 3 and Grade 4adverse events occurred in 41 cases. This is the first to report results from the prospectively 7-year follow-up of Chinesepatients infected HIV taking HAART. It demonstrates that two NRTIs and one NNRTI regimens may persistentlysuppress HIV viremia and continuously induce CD4 cell increase, with good safety and tolerance. The majority took firstlineregimens effectively. 19.2% changed to other first-line drug due to drug-related side effects, 10.2% switched tosecond-line regimens due to viral resistance. Some discontinued or got virological failure because of poor compliance.

Once-a-Day (QD) vs Twice-Daily (BID) Nevirapine as Simplification in PITreated Patients After 2 mos. of BID Induction by Renato Maserati, Micaela Brandolini, Annamaria Cattelan, Anna Orani, Laura Sighinolfi, Massimo Andreoni, Alessia Uglietti, Giovanni Guaraldi, Giovanni Sotgiu (166-173).
To assess the efficacy and the tolerability of once-daily (QD) versus twice-daily (BID) nevirapine (NVP)-basedhighly active antiretroviral therapy (HAART) in virologically suppressed, HIV-positive patients switched from proteaseinhibitor (PI)-based HAART.Eligible patients were enrolled in the multicenter trial if HIV RNA levels were <50 copies/mL for ≥6 months prior.Patients were switched from a PI to NVP 200 mg BID for 2 months, and then randomized to continue with that regimen(group A) or NVP 400 mg QD (group B) for a further 10 months. Virological efficacy (primary endpoint) andtolerability/toxicity were evaluated according to an intention-to-treat analysis.A total of 126 patients (63 per group) were enrolled. Withdrawals from the study (any reason) numbered 15 in group Aand 14 in B, virological failures numbered 5 and 2, respectively, and there were 4 cases of adverse events in each group(all p = NS). Mean alanine aminotransaminase (ALT) and gamma-glutamyl transpeptidase (γ-GT) level increases weresignificant for the whole cohort (33.2±22.9 to 43.3±29.1, p ‹ 0.001; 57.3±72 to 109±131 U/L, p ‹ 0.0002, respectively),but there were no differences between the two groups.Apparently, no significant differences between the QD and BID NVP groups were found, in terms of virological failuresor tolerability/toxicity. The switch to NVP may be safely pursued with a QD schedule.

Neutral Actions of Raltegravir on Adipogenesis, Glucose Metabolism and Lipolysis in 3T3-L1 Adipocytes by Patricia Perez-Matute, Laura Perez-Martinez, Jose R. Blanco, Jose A. Oteo (174-179).
Raltegravir (RAL) has been shown to be virologically effective in both treatment naive and triple class resistantpatients. A more favourable metabolic profile associated with RAL in comparison with other antiretroviral drugs has alsobeen observed. The aim of this study was to investigate the molecular mechanisms that could explain the lack of toxicityof this drug in metabolism. Thus, the effects of RAL on adipogenesis and adipocyte metabolism were analyzed using 3T3-L1 cells, a very adequate and convenient cell culture model for the investigation of adipose differentiation andmetabolism. The effects of RAL on adipogenesis were evaluated by the Oil Red O staining after 8 days of induction ofdifferentiation. Several adipogenic genes (C/EBPα, PPARγ, Pref-1 and AP2) were analyzed by Real-Time PCR. Fullydifferentiated adipocytes were also incubated with RAL for 24 hours and glucose utilization, lactate production andglycerol release were analyzed. Thus, minimal effects of RAL on murine adipocyte differentiation were observed. Basalglucose uptake and lactate production were not affected by RAL at any of the concentrations used. No effects were alsofound on the percentage of glucose that is metabolized to lactate. Lipolysis was only slightly inhibited by Raltegravir (-10%) at the highest concentration used (50 µM), while no effects were observed with lower doses. Our results suggest thatthe absence of significant actions of RAL on adipogenesis and glucose and lipid metabolism in adipocytes could explain,at least in part, the neutral metabolic effects of RAL in clinical studies.

Cervical Squamous Intraepithelial Lesions Among HIV-Positive Women on Antiretroviral Therapy in Kenya by Kevin P. McKenzie, Robyn K. Rogers, Julia W. Njoroge, Grace John-Stewart, Barbra A. Richardson, Nelly R. Mugo, Hugo De Vuyst, Ritesh N. Pamnani, Farzana S. Rana, Danson Warui, Michael H. Chung (180-185).
Background: The prevalence of cervical squamous intraepithelial lesions (SIL) among HIV-infected women onantiretroviral therapy in sub-Saharan Africa has not been well described.Methods: HIV-infected women enrolled in an HIV treatment clinic in Nairobi, Kenya were offered free cervical screeningwith Papanicolaou (Pap) smear testing if they were 30 to 39 years of age and on antiretroviral therapy. Women with SILwere compared to those without SIL with univariate analyses and logistic regression.Results: Of 595 eligible women, 267 accepted Pap testing and had available cytology results, of whom 258 (97%) were ona non-nucleoside reverse transcriptase inhibitor (NNRTI) based regimen. Median duration of antiretroviral therapy was 13months [interquartile range (IQR), 8-19]. Abnormal cytology was found in 123 women (46%) with 70 women (26%)having low grade squamous intraepithelial lesions (LSIL), 22 (8%) high grade squamous intraepithelial lesions (HSIL), 30(11%) atypical squamous cells of unknown significance (ASCUS) and 1 (0.4%) atypical glandular cells (AGC). Womenwith SIL had lower median CD4 cell count (239 vs 287 cells/mm3; P=0.02), lower income (<70 USD per month: 57% vs38%; P=0.01), and less regular condom use (24% vs 40%; P=0.02) compared to those with no SIL. Duration and type ofantiretroviral regimen were not significantly associated with SIL.Conclusion: SIL is prevalent among women on antiretroviral therapy and is associated with immunosuppression, lowincome, and less frequent condom use. Cervical cancer screening and counseling on condom use should be routinelyoffered to HIV-infected women in antiretroviral treatment clinics in Africa.

Liver-Related Factors Associated with Low Vitamin D Levels in HIV and HIV/HCV Coinfected Patients and Comparison to General Population by Laura Milazzo, Cristina Mazzali, Giovanna Bestetti, Erika Longhi, Antonella Foschi, Anita Viola, Tarcisio Vago, Massimo Galli, Carlo Parravicini, Spinello Antinori (186-193).
Objectives: Low 25-Hydroxyvitamin D (25[OH]D) was associated with severe fibrosis and low sustainedvirological response (SVR) after interferon (IFN)-based therapy in chronic hepatitis C. Furthermore, hypovitaminosis Dwas reported in HIV-infected individuals, but its role in liver disease progression in HIV/HCV coinfection is unknown.Methods: 25(OH)D was retrospectively measured in 237 HIV-infected patients (93 with HCV coinfection) and 76 healthycontrols. Multivariate analysis included season, immuno-virological data, combined antiretroviral therapy (cART) and, ina subgroup of 51 HIV/HCV-genotype 1 coinfected patients, factors influencing SVR to pegylated-IFN and ribavirin. In agroup of 20 patients, liver expression of cytochrome (CY)-P27A1 and CYP2R1, 25-hydroxylating enzymes, was assessedby immunohistochemistry.Results: Median 25(OH)D levels were 23.4 (interquartile range 16.7-33.7) ng/mL in the HIV-infected population and 24ng/mL (18.3-29.5) in healthy controls (p=0.9). At multiple regression analysis, only winter/spring measurementscorrelated with lower 25(OH)D levels. No correlation with HCV coinfection, nor with cART regimens was found. Low25(OH)D was independently associated with advanced fibrosis in HIV/HCV coinfected patients (p=0.023), whereas noassociation emerged with SVR to IFN-based therapy. CYP27A1 and CYP2R1 expression was associated neither with25(OH)D serum levels nor with HCV-infection, liver histology, or cART.Conclusions: In our experience, despite the high prevalence of 25(OH)D insufficiency, HIV and HCV-infection did notseem to influence vitamin D status. The role of HIV, HCV and cART on hypovitaminosis D needs further validation inlarger cohorts that account for the vitamin levels in general populations and for seasonal and regional variability.

Background: Canada's international response to HIV may be under threat given CIDA's new aid priorities thatappear to exclude health. Drivers of this recent priority shift have included the influence of global aid trends among publicsector donors and changes within the global HIV milieu itself. However, this is not the first time Canada has shifted inresponse to these two global trends. The era from 2000-2004 also witnessed dramatic changes in both the HIV field and inglobal thinking around international aid. As such, this article presents an evaluation of the Government of Canada'sinternational response to HIV during the first era of transition (2000-2004) in order to derive lessons for decision-makingaround HIV in the current climate of change.Methods: In-depth, semi-structured interviews were conducted with 23 key informants with expertise regarding Canada'sinternational response to HIV over time. Analysis involved multiple readings of transcripts to identify descriptive codesand establish intimacy with the data. Descriptive codes were then collapsed into thematic categories using a process ofinductive reasoning.Results: Canada's international response to HIV was perceived to be exemplary at times (e.g. seminal funding to WHO's3-by-5 strategy), but also inconsistent (e.g., underutilized technical assistance capacity) and non-strategic (e.g.,contradiction between investing in training health providers while poaching professionals to bolster Canada's workforce).Conclusions: Lessons from the 2000-2004 era of transition focus on strategic investments, the inextricable connectionbetween HIV and development, and strategy coherence. These results highlight that it is more constructive to ensure thatCanadian development responses in all areas engage with both the upstream drivers of HIV as well as the impacts of theepidemic itself in order to achieve the greatest results from international investment and the most effective contributionsto the lives of the people that these endeavours seek to support.