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Natural orthopoxvirus outbreaks such as vaccinia, cowpox, cattlepox and buffalopox continue to cause morbidity in the human population. Monkeypox virus remains a significant agent of morbidity and mortality in Africa. Furthermore, monkeypox virus's broad host-range and expanding environs make it of particular concern as an emerging human pathogen. Monkeypox virus and variola virus (the etiological agent of smallpox) are both potential agents of bioterrorism. The first line response to orthopoxvirus disease is through vaccination with first-generation and second-generation vaccines, such as Dryvax and ACAM2000. Although these vaccines provide excellent protection, their widespread use is impeded by the high level of adverse events associated with vaccination using live, attenuated virus. It is possible that vaccines could be used in combination with antiviral drugs to reduce the incidence and severity of vaccine-associated adverse events, or as a preventive in individuals with uncertain exposure status or contraindication to vaccination. We have used the intranasal mousepox (ectromelia) model to evaluate the efficacy of vaccination with Dryvax or ACAM2000 in conjunction with treatment using the broad spectrum antiviral, brincidofovir (BCV, CMX001). We found that co-treatment with BCV reduced the severity of vaccination-associated lesion development. Although the immune response to vaccination was quantifiably attenuated, vaccination combined with BCV treatment did not alter the development of full protective immunity, even when administered two days following ectromelia challenge. Studies with a non-replicating vaccine, ACAM3000 (MVA), confirmed that BCV's mechanism of attenuating the immune response following vaccination with live virus was, as expected, by limiting viral replication and not through inhibition of the immune system. These studies suggest that, in the setting of post-exposure prophylaxis, co-administration of BCV with vaccination should be considered a first response to a smallpox emergency in subjects of uncertain exposure status or as a means of reduction of the incidence and severity of vaccine-associated adverse events.

CMX001 is an orally available lipid acyclic nucleotide phosphonate that delivers high intracellular levels of cidofovir (CDV)-diphosphate and exhibits enhanced in vitro antiviral activity against a wide range of double-stranded DNA viruses, including cytomegalovirus (CMV). Mutations in the DNA polymerase of CMV that impart resistance to CDV also render the virus resistant to CMX001. Here, we report a novel resistance mutation that arose under the selective pressure of CMX001. The wild-type CMV strain AD169 was propagated in human foreskin fibroblasts under increasing concentrations of CMX001 over 10 months, and the resulting strain (named CMX001 R ) was less susceptible to CDV and CMX001 in a plaque reduction assay. Genotypic analysis of virus strain CMX001 R via conventional sequencing of the genes encoding the CMV DNA polymerase ( UL54 ) and UL97 kinase ( UL97 ) demonstrated one mutation that changed the wild-type aspartate to glutamate at position 542 in UL54 . A recombinant virus with this novel D542E mutation was generated via bacterial artificial chromosome-mediated marker transfer experiments. Subsequent phenotypic resistance analysis of the D542E mutant demonstrated reductions in susceptibility of greater than 10-fold to CMX001 and CDV, but no resistance to foscarnet (FOS) or ganciclovir (GCV). Analysis of replicative fitness showed that both strain CMX001 R and the D542E mutant viruses demonstrated a smaller plaque phenotype and slower replication kinetics than their respective parent viruses. These data describe the first resistance mutation generated under the selective pressure of CMX001 and suggest that CMX001 may have a unique resistance profile associated with reduced viral replication and maintenance of sensitivity to FOS and GCV.

Eight in vitro selection experiments under brincidofovir pressure elicited the known cytomegalovirus DNA polymerase amino acid substitutions N408K and V812L and the novel exonuclease domain substitutions D413Y, E303D, and E303G, which conferred ganciclovir and cidofovir resistance with 6- to 11-fold resistance to brincidofovir or 17-fold when E303G was combined with V812L. The new exonuclease domain I resistance mutations selected under brincidofovir pressure add to the single instance previously reported and show the expected patterns of cross-resistance.

Although acyclovir (ACV) has proven to be of value in the therapy of certain herpes simplex virus (HSV) infections, there is a need for more effective therapies, particularly for serious infections in neonates and immunocompromised individuals, where resistance to this drug can be problematic. CMX001 is an orally bioavailable lipid conjugate of cidofovir that is substantially less nephrotoxic than the parent drug and has excellent antiviral activity against all the human herpesviruses. This compound retains full antiviral activity against ACV-resistant laboratory and clinical isolates. The combined efficacy of CMX001 and ACV was evaluated in a new real-time PCR combination assay, which demonstrated that the combination synergistically inhibited the replication of HSV in cell culture. This was also confirmed in murine models of HSV infection, where the combined therapy with these two drugs synergistically reduced mortality. These results suggest that CMX001 may be effective in the treatment of ACV-resistant HSV infections and as an adjunct therapy in individuals with suboptimal responses to ACV. Effective therapies are available for the management of both genital herpes (23) and herpes labialis (1). Acyclovir (ACV) can inhibit the replication of both herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) (6), and its oral prodrug, valacyclovir, is well tolerated and suitable for long-term suppressive therapy (38). Prophylactic therapy in individuals with frequent or severe symptoms can reduce the frequency of lesions by 80% to 95% (19) and can also reduce significantly, although not eliminate, asymptomatic virus shedding (8, 11) and transmission of the virus (5). Resistance to the drug is observed infrequently in the general population, even after long-term prophylactic therapy (38), and resistance occurs in immunocompetent individuals at a very low frequency (0.5%) (17). However, resistance to the drug has been reported to arise in up to 7% of immunocompromised hosts (38). Drug-resistant isolates selected by ACV therapy are also generally crossresistant to famciclovir, and secondary therapies such as cidofovir (CDV) and foscarnet are associated with significant toxicities. Thus, there is a clear need for improved therapies for herpes infection in immunocompromised hosts (21, 42).

Brincidofovir (BCV) is the 3-hexadecyloxy-1-propanol (HDP) lipid conjugate of the acyclic nucleoside phosphonate cidofovir (CDV). BCV has established broad-spectrum activity against double-stranded DNA (dsDNA) viruses; however, its activity against RNA viruses has been less thoroughly evaluated. Here, we report that BCV inhibited infection of Ebola virus in multiple human cell lines. Unlike the mechanism of action for BCV against cytomegalovirus and other dsDNA viruses, phosphorylation of CDV to the diphosphate form appeared unnecessary. Instead, antiviral activity required the lipid moiety and in vitro activity against EBOV was observed for several HDP-nucleotide conjugates.

We retrospectively studied the effect of the lamivudine-induced reverse transcription mutation M184V on selection of thymidine analog mutations (TAMs) in HIV subtype C-infected children and on clinical outcome.We genotyped 135 blood samples from 55 children. TAMs accumulation, viral load and clinical outcome were compared in children maintained on zidovudine/stavudine + lamivudine + protease inhibitor/nonnucleoside reverse transcriptase inhibitor (PI/NNRTI) despite loss of viral suppression and in children treated with, or switched to, other nucleoside reverse transcriptase inhibitors (NRTIs). Drug susceptibility and replication capacity of selected samples were measured.M184V developed in 18 of 22 of children who had received only zidovudine/stavudine + lamivudine + PI/NNRTI during a mean of 23.2 +/- 3.2 months versus in 3 of 14 children treated with other drugs and/or having multiple regimen changes (P = 0.001). TAMs appeared, respectively, in 2 of 22 versus 12 of 14 (P0.0001). The 2 groups did not differ significantly in baseline HIV-RNA or CD4 count, sampling time, and follow-up period. In M184V-containing samples, we found large reductions in susceptibility to lamivudine and emtricitabine but not to other NRTIs. When T215Y was present without M184V, susceptibility to zidovudine was reduced 8-fold. When both M184V + T215Y occurred, susceptibility to zidovudine was substantially increased. Average inhibition concentration 50 values were similar to those documented in the Stanford database for subtype B HIV with these mutation patterns.Maintaining a thymidine analog + lamivudine-based regimen reduced accumulation of TAMs and increased zidovudine susceptibility. This is likely the result of an increased susceptibility to thymidine analog (zidovudine) in the context of M184V documented here for the first time in subtype C-infected children. This retrospective study supports the strategy of maintaining lamivudine-containing therapy in subtype C-infected children. This strategy may be beneficially applied in the treatment of children in Africa, where thymidine analog + lamivudine-based regimen became available recently but further options are limited.

Background. Maximum suppression of virus replication is often not achievable for persons infected with multidrug-resistant human immunodeficiency virus type 1 (HIV-1). Available data suggest that lamivudine contributes to partial viral suppression, despite the presence of M 184V mutations and high-level phenotypic lamivudine resistance. Methods. Selective lamivudine withdrawal was studied in 6 subjects who had incomplete viral suppression during antiretroviral treatment for multidrug-resistant HIV-1 infection. Results. Plasma levels of HIV-1 RNA increased to 0.5 log 10 copies/mL above baseline 6 weeks after the withdrawal of lamivudine treatment (P = .04), even though reversion of lamivudine resistance was not yet detected. Early increases in plasma levels of HIV-1 RNA after lamivudine withdrawal were associated with the presence of the T215Y/F mutation and broad phenotypic resistance to nucleoside reverse-transcriptase inhibitors at baseline. Genotypic and phenotypic reversion of lamivudine resistance was detected in 4 subjects 8-14 weeks after withdrawal of lamivudine therapy. The duration of lamivudine withdrawal ranged from 8 to 22 weeks; all subjects resumed lamivudine treatment. Plasma levels of HIV-1 RNA were 0.6 log 10 copies/mL above baseline (P = .03) when lamivudine therapy was resumed. After the resumption of lamivudine treatment, plasma HIV RNA levels decreased to baseline levels in 3 subjects but remained elevated in 3 subjects who had evolution of increased antiretroviral drug resistance during the period of lamivudine withdrawal. Safety concerns raised by this latter finding led to permanent closure of the study. Conclusions. In select cases of multidrug-resistant HIV-1 infection, lamivudine contributes to suppression of HIV-1 replication, despite the presence of M184V mutations and lamivudine resistance.

Background: The ESS40013 study tested 4-drug induction followed by 3-drug maintenance as initial antiretroviral therapy (ART) to reduce HIV RNA rapidly and then to simplify to an effective yet more convenient and tolerable regimen. Methods: Four hundred forty-eight antiretroviral-naive adults were treated with abacavir/lamivudine/zidovudine (ABC/3TC/ZDV) and efavirenz (EFV) for the 48-week induction phase. Two hundred eighty-two patients were randomized in a 1:1 ratio to continue ABC/3TC/ZDV + EFV or to simplify to ABC/3TC/ZDV for the 48-week maintenance phase. Results: The baseline median HIV RNA level and CD4 cell count were 5.08 logic copies/mL (56% ≥ 100,000 copies/mL) and 210 cells/mm 3 (48%

Background. The pathogenesis of HIV encephalopathy is poorly understood especially in children. Studies suggest that HIV replication and the release of proinflammatory mediators in the central nervous system contribute to the pathogenesis of HIV dementia in adults. Methods. Cerebrospinal fluid (CSF) and plasma samples from 23 HIV-infected children were longitudinally analyzed at Weeks 0, 8, 16 and 48 for HIV RNA and concentrations of the following proinflammatory mediators: monocyte chemotactic protein-1 (MCP-1), tumor necrosis factor-alpha, regulated upon activation, normal T cell expressed and secreted (RANTES), macrophage-inflammatory protein (MIP)-1-alpha, MIP-1-beta and matrix metalloproteinase-9 (MMP-9). Results. All 23 children had detectable concentrations of MCP-1 in the CSF at all time points evaluated. However, of the remaining of proinflammatory mediators measured in CSF at baseline, only a few children had detectable concentrations: tumor necrosis factor-alpha, n = 1; RANTES, n = 5; MMP-9, n = 9; MIP-1-alpha and MIP-1-beta, n = 0. A reduction from baseline to Week 48 was observed in CSF concentrations of MCP-1 and, among children with detectable values, MMP-9, which paralleled declines in CSF HIV RNA. Conclusion. These results suggest that MCP-1 and MMP-9 may be involved in the pathogenesis of central nervous system disease in HIV-infected children.

Objective To assess the relative in vitro potency of the antiviral agents emtricitabine (FTC), lamivudine (3TC), and zidovudine (ZDV) in peripheral blood mononuclear cells (PBMCs), monocyte-derived macrophages, and MT-4 cells infected with HIV-1. DESIGN In vitro evaluation of the test compounds against M-tropic or T-tropic laboratory strains of HIV-1 and against clinical HIV-1 isolates from antiretroviral therapy-naive subjects using PBMCs, monocyte-derived macrophages, and MT-4 cells. Methods Standard methods for assessing antiviral potency based on 50% inhibitory concentrations using propidium iodide staining of host cell DNA to assess cytopathic effects or measurement of HIV-1 reverse transcriptase activity to assess inhibition of viral replication. Results There were no significant differences in potency between 3TC and FTC in assays with HIV-1(IIIB)-infected PBMCs or HIV-1(Ba-L) -infected monocyte-derived macrophages, which are primary cell types for HIV-1 infection in vivo. In agreement with earlier reports, FTC was approximately fourfold more active than 3TC in assays in the transformed T-cell line MT-4 infected with HIV-(1IIIB), whereas ZDV was more active than FTC. 3TC, FTC, and ZDV were equally active against a panel of eight primary HIV-1 isolates from antiretroviral-naive subjects in PBMCs. These results demonstrate the in vitro similarity of 3TC and FTC activity in primary cells. The variability in potency depending on cell types and viral strains underscores our observation that antiviral effects in vitro are not reliable predictors of in vivo clinical activity.

Mycophenolic acid (MPA) enhances the in vitro activity of abacavir (ABC) and other nucleoside analog reverse transcriptase inhibitors (NRTIs) against sensitive and NRTI-resistant HIV-1. This may occur via depletion of intracellular deoxyguanosine triphosphate (dGTP). Mycophenolate mofetil (MMF) 500 mg twice daily was added as a single agent to the antiretroviral regimens of five patients failing maximal available therapy. Therapy included ABC, and in most cases didanosine (DDI) and tenofovir (TDF). At entry, mean plasma HIV-1 RNA (VL) was 5.02 log copies/mL (median 4.78, range 4.71-5.63) and mean CD4 count was 106/microL (median 117, range 11-174). MMF was well tolerated. CD4 cell counts did not change significantly from baseline for up to 60 weeks of follow-up. Three of five subjects had VL declines of >0.5 log copies/mL immediately after adding MMF; a fourth subject had a sustained decline of >0.5 log copies/mL after week 8. Declines of >0.5 log copies/mL were lost in two patients at 6 and 8 weeks, and persisted in two patients at 36 and 60 weeks of follow-up, respectively. An increase in the ratio of carbovir triphosphate (CBV-TP), the active antiviral metabolite of ABC, to dGTP was documented in 3 of 4 subjects in temporal association with decreased VL. Trough plasma MPA levels ranged from 0.26-1.67 microg/mL; peak levels 90 minutes after dosing from 1.20-7.77 microg/mL. AUC of MPA appeared little changed when measured over 28 weeks of therapy. Declines in VL were observed in association with measurable changes in the CBV-TP/dGTP ratio in some patients, whereas MPA AUC was below the 30-60 microg*hr/mL range targeted in organ transplantation. The possibility that MMF may enhance the effect of selected NRTIs and be tolerated in late stage HIV disease deserves careful randomized study.

Macrophages express the chemokine receptor CCR-5, a coreceptor for human immunodeficiency virus (HIV) entry. This receptor is ligated by beta chemokines, which influence HIV type 1 (HIV-1) replication in CCR-5-bearing cells in vitro and could influence the course of infection in the central nervous system. Cerebrospinal fluid (CSF) samples from 73 HIV-infected men were assayed for macrophage inflammatory protein-1 alpha (MIP-1alpha), MIP-1beta, and regulated upon activation, normal T cell expressed and secreted (RANTES). Distributions of all three were positively skewed. CSF chemokine concentrations were correlated with each other and were higher in demented patients. In a multivariate analysis, demented subjects were more likely to have detectable CSF MIP-1alpha, elevated CSF HIV RNA levels, and lower CD4+ cell counts. However, among those with detectable CSF MIP-1alpha, levels were lower in demented patients. CSF beta chemokine elevation is consistent with the macrophage activation known to occur in dementia and with studies of beta chemokine mRNA expression in the brain. Low, but detectable, levels of CSF MIP-1alpha were strongly associated with dementia, suggesting that higher levels may have neuroprotective effects.

Brincidofovir (CMX001), a lipid conjugate of the acyclic nucleotide phosphonate cidofovir, is under development for smallpox treatment using “the Animal Rule,” established by the FDA in 2002. Brincidofovir reduces mortality caused by orthopoxvirus infection in animal models. Compared to cidofovir, brincidofovir has increased potency, is administered orally, and shows no evidence of nephrotoxicity. Here we report that the brincidofovir half-maximal effective concentration (EC 50 ) against five variola virus strains in vitro averaged 0.11 μM and that brincidofovir was therefore nearly 100-fold more potent than cidofovir.

Adenovirus infection is a serious and often fatal complication in hematopoietic stem cell transplant patients. There are currently no FDA-approved therapies for adenovirus infection, with only anecdotal, off-label uses described for a variety of anti-viral agents or immune therapies. We report the first case of successful eradication of disseminated adenovirus infection by the novel antiviral agent CMX001 in a severely immunocompromised pediatric stem cell transplant recipient following failure to respond to intravenous cidofovir. Complete clinical and virologic response was documented; virologic and pharmacokinetic data are reported. CMX001 is a promising new oral antiviral agent under development for the prophylaxis and treatment of severe infections caused by double-stranded DNA viruses including adenovirus in immunocompromised patients.

CMX157 is a lipid (1-0-hexadecyloxypropyl) conjugate of the acyclic nucleotide analog tenofovir (TFV) with activity against both wild-type and antiretroviral drug-resistant HIV strains, including multidrug nucleoside/nucleotide analog-resistant viruses. CMX157 was consistently >300-fold more active than tenofovir against multiple viruses in several different cell systems. CMX157 was active against all major subtypes of HIV-1 and HIV-2 in fresh human peripheral blood mononuclear cells (PBMCs) and against all HIV-1 strains evaluated in monocyte-derived macrophages, with 50% effective concentrations (EC50s) ranging between 0.20 and 7.2 nM. The lower CMX157 EC50s can be attributed to better cellular uptake of CMX157, resulting in higher intracellular levels of the active antiviral anabolite, TFV-diphosphate (TFV-PP), inside target cells. CMX157 produced >30-fold higher levels of TFV-PP in human PBMCs exposed to physiologically relevant concentrations of the compounds than did TFV. Unlike conventional prodrugs, including TFV disoproxil fumarate (Viread), CMX157 remains intact in plasma, facilitating uptake by target cells and decreasing relative systemic exposure to TFV. There was no detectable antagonism with CMX157 in combination with any marketed antiretroviral drug, and it possessed an excellentin vitrocytotoxicity profile. CMX157 is a promising clinical candidate to treat wild-type and antiretroviral drug-resistant HIV, including strains that fail to respond to all currently available nucleoside/nucleotide reverse transcriptase inhibitors.

Thymidine-sparing triple-nucleoside regimens have exhibited poor virologic response despite apparent phenotypic susceptibility to 2 of 3 regimen components at early time points. Phenotypic resistance masking by wild-type virus may explain this discrepancy.Consistent with this notion were (1) the presence of low level nucleoside reverse-transcriptase inhibitor-resistant human immunodeficiency virus in subjects receiving failing first-line regimens consisting of tenofovir (TDF), abacavir (ABC), and lamivudine (3TC); (2) lower fold resistance associated with mixtures versus mutants in a clinical-isolate database; and (3) dose dependent changes in susceptibility to ABC, 3TC, TDF, and didanosine on titration of K65R and/or M184V with wild-type virus. These findings underscore the limitations of stand-alone phenotypic susceptibility measures and emphasize the importance of complementary and/or more sensitive techniques.

Background Transmitted HIV-1 drug resistance can compromise initial antiretroviral therapy (ART); therefore, its detection is important for patient management. The absence of drug-associated selection pressure in treatment-naïve persons can cause drug-resistant viruses to decline to levels undetectable by conventional bulk sequencing (minority drug-resistant variants). We used sensitive and simple tests to investigate evidence of transmitted drug resistance in antiretroviral drug-naïve persons and assess the clinical implications of minority drug-resistant variants. Methods and Findings We performed a cross-sectional analysis of transmitted HIV-1 drug resistance and a case-control study of the impact of minority drug resistance on treatment response. For the cross-sectional analysis, we examined viral RNA from newly diagnosed ART-naïve persons in the US and Canada who had no detectable (wild type, n = 205) or one or more resistance-related mutations (n = 303) by conventional sequencing. Eight validated real-time PCR-based assays were used to test for minority drug resistance mutations (protease L90M and reverse transcriptase M41L, K70R, K103N, Y181C, M184V, and T215F/Y) above naturally occurring frequencies. The sensitive real-time PCR testing identified one to three minority drug resistance mutation(s) in 34/205 (17%) newly diagnosed persons who had wild-type virus by conventional genotyping; four (2%) individuals had mutations associated with resistance to two drug classes. Among 30/303 (10%) samples with bulk genotype resistance mutations we found at least one minority variant with a different drug resistance mutation. For the case-control study, we assessed the impact of three treatment-relevant drug resistance mutations at baseline from a separate group of 316 previously ART-naïve persons with no evidence of drug resistance on bulk genotype testing who were placed on efavirenz-based regimens. We found that 7/95 (7%) persons who experienced virologic failure had minority drug resistance mutations at baseline; however, minority resistance was found in only 2/221 (0.9%) treatment successes (Fisher exact test, p = 0.0038). Conclusions These data suggest that a considerable proportion of transmitted HIV-1 drug resistance is undetected by conventional genotyping and that minority mutations can have clinical consequences. With no treatment history to help guide therapies for drug-naïve persons, the findings suggest an important role for sensitive baseline drug resistance testing., Using real-time PCR to detect HIV resistance mutations present at low levels, Jeffrey Johnson and colleagues investigate prevalence and clinical implications of minority transmitted mutations., Editors' Summary Background Since the mid-1990s, several powerful antiretroviral drug combinations have been developed that have greatly improved the prognosis of HIV infection. All antiretroviral therapy (ART) regimens combine drugs that act against HIV in different ways (so-called different drug classes). Multiple drugs are necessary because HIV continually accumulates random changes (mutations) in its genetic material (genome). Some of these mutations make HIV resistant to individual antiretroviral drugs, so a mixture of drugs is needed to keep the virus in check. However, the efficacy of ART (which itself selects for drug-resistant variants by giving them a growth advantage over drug-sensitive variants) is substantially reduced when these variants account for more than about 20% of the viruses in an infected person. This level of variant virus can be detected in blood samples with a technique called bulk sequencing. In North America and Europe, where ART has been widely used for many years, around 20% of HIV-infected people who have taken ART themselves develop this level of drug-resistant virus, which can be transmitted by the same routes as nonresistant HIV (typically unprotected sexual intercourse or needle sharing). In such cases, the person acquiring drug-resistant HIV may experience treatment failure when drugs later fail to work against the resistant virus. In these countries, therefore, resistance testing by bulk sequencing is done routinely before ART is initiated to decide which antiviral drugs are likely to be effective. Why Was This Study Done? Several years usually elapse between the time a person becomes infected with HIV and the time he or she starts ART. During this time, the absence of selection pressure from antiviral drugs means that transmitted drug-resistant variants tend to decline to levels undetectable by bulk sequencing. These “minority drug-resistant variants” can be detected using other more sensitive tests but it is not known what proportion of HIV-infected people who have never taken ART carry minority drug-resistant variants (the “prevalence” of these variants). It is also unknown whether the presence of minority drug-resistant variants reduces the success of ART. In this paper, the researchers first report a “cross-sectional” study in North America using a sensitive assay to determine the prevalence of minority drug-resistant viruses among HIV-infected people who had never received ART. They then investigate whether minority drug-resistant variants have any impact on the effectiveness of ART in a “case-control” study. What Did the Researchers Do and Find? In their cross-sectional study, the researchers used a highly sensitive test for detecting mutations (called a real-time PCR-based assay) to look for low levels of viruses carrying any of eight major drug-resistance mutations in people with newly diagnosed HIV infection who reported no prior treatment with ART. Seventeen percent of the people who had only wild-type (nonmutated) virus by bulk sequencing (205 participants) were found, in fact, to carry low levels of virus variants with 1–3 drug-resistance mutations; 2% of them carried viruses resistant to two different drug classes (called multi-drug resistance). Among the people with resistance mutations detected by bulk sequencing (303 participants), 10% had at least one additional minority drug-resistant variant, often a viral variant that was resistant to a drug class different from that detected by bulk sequencing. In the case-control study, the researchers used their sensitive assays to measure the levels of viruses containing any of the three most common drug resistance mutations likely to affect viral responses to the antiretroviral drugs efavirenz and lamivudine in 316 people just before they started their first HIV treatment, which included these drugs. Of people for whom ART failed, 7% were infected with minority drug-resistant virus variants at baseline compared with only 0.9% of people for whom ART worked; this difference was statistically significant. What Do These Findings Mean? The findings of the cross-sectional study indicate that conventional bulk sequencing fails to detect a large proportion of transmitted HIV drug resistance and suggest that the transmission of drug-resistant variants from infectious ART-experienced people to ART-naïve individuals might not be uncommon. The findings of the case-control study suggest that the minority drug-resistant HIV variants may have clinical consequences. That is, the presence of such variants in individuals who have not previously taken ART may reduce the efficacy of some ART regimens. However, the number of participants meeting the criteria for analysis in the cross-sectional study was limited, and the association between minority resistance and treatment failure may have been influenced by other factors. Taken together, these findings suggest that, to ensure that first-line ART is as effective as possible, greater efforts should be made to prevent HIV transmission, whether from ART-experienced or ART-naive people. However, because data on minority drug-resistant virus are limited, more studies— particularly with recent populations—are needed before testing for these variants can be considered appropriate in the clinical management of newly diagnosed HIV infection. Additional Information. Please access these Web sites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.0050158. This study is further discussed in a PLoS Medicine Perspective by Steven G. Deeks Information is available from the US National Institute of Allergy and Infectious Diseases on HIV infection and AIDS HIV InSite has comprehensive information on all aspects of HIV/AIDS, including links to fact sheets (in English, French, and Spanish) about antiretrovirals and information on genetic testing for HIV drug resistance NAM, a UK registered charity, provides information about all aspects of HIVand AIDS, including fact sheets on types of HIV drug, drug resistance, and resistance tests (in English, Spanish, French, Portuguese, and Russian) The US Centers for Disease Control and Prevention provides information on HIV/AIDS and on treatment (in English and Spanish)

Objective: To identify optimal first-line therapies based on the rate of virologic success (VS) and the preservation of future treatment options in antiretroviral therapy (ART)-naive subjects. Design: Systematic overview of genotypic resistance mutations from clinical trials of combination ART. Methods: Various sources were searched for studies in ART-naive subjects providing virologic response rates and genotypes from subjects with virologic failure. The International AIDS Society-USA genotypic resistance guidelines were used to calculate regimen resistance cost (RC reg ) and number of active drug (AD) scores for each regimen and to rank the regimens. Results: Intra- and interstudy comparisons showed higher VS rates for nonnucleoside reverse transcriptase inhibitor (NNRTI) regimens (range: 51%-76%) and boosted protease inhibitor (boosted PI) regimens (range: 55%-79%). Boosted PI failures had the lowest RC reg (range: 0.12-0.21) and the highest AD (range: 19.80-20.18) scores. NNRTI failures had higher RC reg (range: 0.00-1.22) and lower AD (range: 16.83-21) scores. Conclusions: NNRTI and boosted PI regimens provide the highest rates of VS in treatment-naive HIV-infected persons. Treatment option scores were higher in subjects who failed boosted PI-containing regimens versus NNRTI-containing regimens, however.

Potential contributors to the high rate of virologic failure observed for tenofovir, abacavir, and lamivudine include a low genetic barrier to resistance for this regimen and antagonistic drug-drug interactions. To examine the second possibility, we tested combinations of abacavir, tenofovir, and lamivudine against wild-type and drug-resistant HIV-1 in vitro using peripheral blood mononuclear cells and MT-4 cells. Antagonistic interactions were not detected for any combination. If the systems examined accurately reflect the in vivo situation, antagonism does not substantially contribute to the poor efficacy of this triple combination.

ObjectiveTo evaluate HIV-1 reverse transcriptase genotypic and phenotypic indicators of resistance to abacavir (ABC) as predictors of ABC antiviral efficacy.DesignThe study was a retrospective, combined analysis of five multicentre trials in which ABC was added as a single agent to background antiretroviral therapy in experienced adults.MethodsBaseline HIV-1 genotype and phenotypic susceptibility to ABC were determined and the association of genotype and phenotype with virological response after addition of ABC was analysed.ResultsOverall, 68% of these therapy-experienced subjects had a virological response (>0.5 log10or ConclusionsVirological response to ABC may be diminished significantly by multiple NRTI-associated mutations and/or by reductions in phenotypic susceptibility to ABC. However, many subjects with baseline samples showing evidence of resistance to NRTIs respond to ABC.

Cerebrospinal fluid (CSF) human immunodeficiency virus (HIV) RNA levels were measured with the Nucleic Acid Sequence-Based Amplification (NASBA) assay to determine the relationship with neurological status; 37 subjects with HIV dementia (HIV-D) were compared with 77 with HIV with minor neurological signs (HIV-MCMD) and 93 neurologically normal HIV-seropositive individuals (HIV-NML). The NASBA assay had a lower limit of detection of 100 copies per milliliter. Mean CSF log HIV RNA levels were significantly higher in those with dementia after adjusting for CD4 count and were correlated with dementia severity. Plasma levels did not distinguish comparably immunosuppressed subjects with or without dementia. CSF and plasma RNA levels were significantly intercorrelated for subjects with CD4 counts200/mm3 and also correlated inversely with CSF beta2-microglobulin. CSF RNA levels were independent of CSF pleocytosis or antiretroviral exposure. Brain RNA levels were consistently higher than CSF but correlated with CSF values for dementia subjects. The NASBA assay can be used reliably to determine HIV RNA levels in CSF, brain, and plasma samples. CSF HIV RNA may be a surrogate marker for brain infection, based on the observed correlation with brain levels. The association between plasma HIV RNA and CSF levels of HIV and beta2-microglobulin suggests that both viral load and CNS immune activation are important determinants of neurological disease.