Humanized mouse models of HIV infection.

Because of the limited tropism of HIV, in vivo modeling of this virus has been almost exclusively limited to other lentiviruses, such as simian immunodeficiency virus, that reproduce many important characteristics of HIV infection. However, there are significant genetic and biological differences among lentiviruses and some HIV-specific interventions are not effective against other lentiviruses in nonhuman hosts. For these reasons, much emphasis has recently been placed on developing alternative animal models that support HIV replication and recapitulate key aspects of HIV infection and pathogenesis in humans.

Tumor suppressor genes FHIT and WWOX are deleted in primary effusion lymphoma (PEL) cell lines.

Primary effusion lymphoma (PEL) is a diffuse-large B-cell lymphoma with poor prognosis. One hundred percent of PELs carry the genome of Kaposi sarcoma-associated herpesvirus and a majority are coinfected with Epstein-Barr virus (EBV). We profiled genomic aberrations in PEL cells using the Affymetrix 6.

HIV-1 Populations in Semen Arise through Multiple Mechanisms.

HIV-1 is present in anatomical compartments and bodily fluids. Most transmissions occur through sexual acts, making virus in semen the proximal source in male donors. We find three distinct relationships in comparing viral RNA populations between blood and semen in men with chronic HIV-1 infection, and we propose that the viral populations in semen arise by multiple mechanisms including: direct import of virus, oligoclonal amplification within the seminal tract, or compartmentalization.

A strongly transdominant mutation in the human immunodeficiency virus type 1 gag gene defines an Achilles heel in the virus life cycle.

The human immunodeficiency virus type 1 (HIV-1) protease (PR) makes five obligatory cleavages in the viral Gag polyprotein precursor. The cleavage events release the virion structural proteins from the precursor and allow the virion to undergo maturation to become infectious. The protease cleavage between the matrix protein (MA) domain and the adjacent capsid protein (CA) domain releases CA from the membrane-anchored MA and allows the N terminus of CA to refold into a structure that facilitates the formation of hexamer arrays that represent the structural unit of the capsid shell.

Cross-sectional characterization of HIV-1 env compartmentalization in cerebrospinal fluid over the full disease course.

Objectives: To characterize HIV-1 env compartmentalization between cerebrospinal fluid (CSF) and peripheral blood plasma over all stages of the HIV-1 disease course, and to determine the relationship between the extent of CSF HIV-1 env compartmentalization and clinical neurologic disease status.

Design: Paired blood plasma and CSF specimens were collected from 66 HIV-infected patients cross-sectionally representing all major clinical stages relating to HIV-associated neurologic disease, including primary infection, asymptomatic chronic infection, chronic infection with minor global impairment, and immune deficiency with HIV-associated dementia.

Methods: Heteroduplex tracking assays and bulk sequence analysis targeting the V1/V2, C2-V3, and V4/V5 regions of env were performed to characterize the genetic makeup of complex HIV-1 populations in the cross-sectional blood plasma and CSF specimens.

Nevirapine resistance in human immunodeficiency virus type 1-positive infants determined using dried blood spots stored for up to six years at room temperature.

Dried blood spots that had been stored ambiently for 3 to 6 years lost approximately 1 log(10) of human immunodeficiency virus type 1 (HIV-1) RNA, but the majority could still be genotyped for resistance. Nevirapine resistance was found in 7/16 (43.5%) HIV-1-positive HIVNET 024 infants at 4 to 6 weeks, but no resistance was found at other time points.

Genotypic susceptibility scores and HIV type 1 RNA responses in treatment-experienced subjects with HIV type 1 infection.

This study compared the role of genotypic susceptibility scores (GSS) as a predictor of virologic response in a group (n = 234) of HIV-infected, protease inhibitor (PI)-experienced subjects. Two scoring methods [discrete genotypic susceptibility score (dGSS) and continuous genotypic susceptibility score (cGSS)] were developed. Each drug in the subject's regimen was given a binary susceptibility score using Stanford inferred drug resistance scores to calculate the dGSS.

Development and characterization of a new single cycle vaccine vector in the simian immunodeficiency virus model system.

We have developed a new single cycle lentiviral vector, SIVsmH4i-SC27.1, as a potential SIV/HIV-1 vaccine candidate. This viral vector is capable of expressing all of the SIV gene products but is limited to one round of infection.

Sequential turnover of human immunodeficiency virus type 1 env throughout the course of infection.

We examined the rates of variant population turnover of the V1-V2 and V4-V5 hypervariable domains of the human immunodeficiency virus type 1 (HIV-1) gp120 molecule in longitudinal plasma samples from 14 men with chronic HIV-1 infection using heteroduplex tracking assays (HTA). Six men had high rates of CD4+ T-cell loss, and eight men had low rates of CD4+ T-cell loss over 2.5 to 8 years of infection.

Genotypic analysis of the protease and reverse transcriptase of HIV type 1 subtype C isolates from antiretroviral drug-naive adults in Malawi.

The protease (PR) and reverse transcriptase (RT) regions of HIV-1 isolates from 21 antiretroviral (ARV)-naive Malawian adults were sequenced and analyzed to determine the prevalence of drug resistance-associated mutations in this population. Phylogenetic analysis confirmed that all isolates grouped with HIV-1 subtype C, the predominant subtype in Malawi. No major mutations associated with resistance to PR inhibitors (PIs), nucleoside RT inhibitors (NRTIs), or nonnucleoside RT inhibitors (NNRTIs) were found.

Increased human immunodeficiency virus type 1 (HIV-1) env compartmentalization in the presence of HIV-1-associated dementia.

The human immunodeficiency virus type 1 (HIV-1) surface Env protein has been implicated in the development of HIV-1-associated dementia (HAD). HIV-1 env diversity was analyzed by heteroduplex tracking assay in 27 infected subjects with various neurological statuses. env compartmentalization between the blood and cerebral spinal fluid (CSF) was apparent with all neurological categories.

Effect of a protease inhibitor-induced genetic bottleneck on human immunodeficiency virus type 1 env gene populations.

The initiation of drug therapy or the addition of a new drug to preexisting therapy can have a significant impact on human immunodeficiency virus type 1 (HIV-1) populations within a person. Drug therapy directed at reverse transcriptase and protease can result in dramatic decreases in virus load, causing a contraction in the virus population that represents a potential genetic bottleneck as a subset of virus with genomes carrying resistance mutations repopulate the host. While this bottleneck exerts an effect directly on the region that is being targeted by the drugs, it also affects other regions of the viral genome.

HIV-1 populations in blood and breast milk are similar.

Mother-to-child transmission (MTCT) of human immunodeficiency virus type 1 (HIV-1) through breast milk is a significant mechanism of infection in many regions of the world. We compared the HIV-1 populations in paired blood and breast milk samples using a heteroduplex tracking assay (HTA) for the V1/V2 regions of env (V1/V2-HTA). V1/V2-HTA patterns were similar in the eight pairs of samples for which adequate template sampling could be demonstrated.

Covariation of amino acid positions in HIV-1 protease.

We have examined patterns of sequence variability for evidence of linked sequence changes in HIV-1 subtype B protease using translated sequences from protease inhibitor (PI) treated and untreated subjects downloaded from the Stanford HIV RT and Protease Sequence Database (http://hivdb.stanford.edu).

Turnover of env variable region 1 and 2 genotypes in subjects with late-stage human immunodeficiency virus type 1 infection.

The env gene of human immunodeficiency virus type 1 (HIV-1) includes some of the most genetically diverse regions of the viral genome, which are called variable regions 1 through 5 (V1 through V5). We have developed a heteroduplex tracking assay to detect changes in variable regions 1 and 2 of env (V1/V2-HTA). Using sequences from two molecular clones as probes, we have studied the nature of longitudinal virus population changes in a cohort of HIV-1-infected subjects.

Selection of high-level resistance to human immunodeficiency virus type 1 protease inhibitors.

Protease inhibitors represent some of the most potent agents available for therapeutic strategies designed to inhibit human immunodeficiency virus type 1 (HIV-1) replication. Under certain circumstances the virus develops resistance to the inhibitor, thereby negating the benefits of this therapy. We have carried out selections for high-level resistance to each of three protease inhibitors (indinavir, ritonavir, and saquinavir) in cell culture.

Replacement of the P1 amino acid of human immunodeficiency virus type 1 Gag processing sites can inhibit or enhance the rate of cleavage by the viral protease.

Processing of the human immunodeficiency virus type 1 (HIV-1) Gag precursor is highly regulated, with differential rates of cleavage at the five major processing sites to give characteristic processing intermediates. We examined the role of the P1 amino acid in determining the rate of cleavage at each of these five sites by using libraries of mutants generated by site-directed mutagenesis. Between 12 and 17 substitution mutants were tested at each P1 position in Gag, using recombinant HIV-1 protease (PR) in an in vitro processing reaction of radiolabeled Gag substrate.

Effects of genital tract inflammation on human immunodeficiency virus type 1 V3 populations in blood and semen.

We have examined cell-free viral populations in the blood plasma and seminal plasma compartments of men infected with subtype C human immunodeficiency virus type 1 (HIV-1) using the V3-specific heteroduplex tracking assay (V3-HTA). We studied two cohorts of subjects who had visited either a sexually transmitted disease (STD) clinic for genital tract inflammation in the form of urethritis (n = 43) or a dermatology clinic (controls, n = 14) in Malawi. We have previously shown that the presence of urethritis is associated with an eightfold increase in virus load in the seminal plasma compartment (M.

Patterns of changes in human immunodeficiency virus type 1 V3 sequence populations late in infection.

We have used a V3-specific heteroduplex tracking assay (V3-HTA) with probes from two different human immunodeficiency virus type 1 (HIV-1) subtypes to examine the extent and pace of HIV-1 evolution late in infection. Twenty-four subjects with advanced HIV-1 infection (CD4(+) T-cell count, <100/microl) and stable viral loads were studied using blood plasma samples collected over a study period of approximately 9 months, during which time most of the subjects were treated with reverse transcriptase inhibitors. The V3-HTA patterns from the first and last time points were evaluated initially to determine the amounts of change in V3 sequence populations, which were primarily changes in abundance in preexisting sequence populations.

Human immunodeficiency virus type-1 protease inhibitors: therapeutic successes and failures, suppression and resistance.

The retroviral protease (PR) is responsible for cleaving precursor proteins that contain the virion structural proteins and enzymes. Highly potent inhibitors of the human immunodeficiency virus type-1 PR have been developed, and to date, five of these inhibitors have been approved for clinical use. These inhibitors bind to the active site of the dimeric PR and represent transition state analogs.

Characterization of V3 sequence heterogeneity in subtype C human immunodeficiency virus type 1 isolates from Malawi: underrepresentation of X4 variants.

We have examined the nature of V3 sequence variability among subtype C human immunodeficiency virus type 1 (HIV-1) sequences from plasma-derived viral RNA present in infected men from Malawi. Sequence variability was assessed by direct sequence analysis of the V3 reverse transcription-PCR products, examination of virus populations by a subtype C V3-specific heteroduplex tracking assay (V3-HTA), and selected sequence analysis of molecular clones derived from the PCR products. Sequence variability in V3 among the subtype C viruses was not associated with the presence of basic amino acid substitutions.

The HIV type 1 protease inhibitor saquinavir can select for multiple mutations that confer increasing resistance.

Previous use of the HIV-1 protease inhibitor saquinavir resulted in the infrequent appearance of mutations in the HIV-1 protease gene associated with resistance. We have examined the ability of saquinavir to select for resistance mutations. In multiple selections of HIV-1 in cell culture with saquinavir, similar patterns of mutations were reproducibly observed and the number of mutations increased with increasing selective pressure.