Clinicopathological and virological characteristics of superficially invasive squamous-cell carcinoma of the anus.

AIM: The clinicopathological and virological characteristics of anal superficially invasive squamous-cell carcinoma (SISCCA) were determined. METHOD: Seventeen patients with a completely excised stage T1N0M0 anal squamous-cell carcinoma (SCC) were included in the study. The tumours were divided into superficially invasive and invasive. Patients with anal high-grade squamous intraepithelial dysplasia, which corresponded to anal intraepithelial neoplasia (AIN) Grades 2 or 3, were used as a control group. Clinicopathological and virological characteristics were investigated. Overall survival and cancer recurrence-free survival were also assessed. RESULTS: Of the 17 patients, 12 (70.5%) were men. Ten (58.8%) were human immunodeficiency virus positive. Seven (41%) patients met the same diagnostic criteria as those recently proposed for anal SISCCA. According to the results obtained using the polymerase chain reaction, human papillomavirus (HPV) 16 was the most commonly detected (94%) type of HPV. Twelve (70.6%) patients with an inadequate surgical margin around the tumour received adjuvant radiotherapy, including the two (11.7%) tumours that locally recurred, one of which was an anal SISCCA. Superficially invasive anal cancers differed from the other T1N0M0 anal carcinomas according to the clinical presentation and the absence of lymph-vascular invasion (LVI). There were no differences in cancer recurrence-free and overall survival rates between the superficially invasive and invasive groups. CONCLUSION: Anal SISCCAs have a low index of clinical suspicion, are associated with an absence of LVI and are linked to high-risk HPV. Prospective studies are needed to define the clinical behaviour of these anal tumours and to determine their best therapeutic strategy.

Dynamics of HIV-1 DNA level in highly antiretroviral-experienced patients receiving raltegravir-based therapy.

To assess dynamics of HIV-1 DNA in highly antiretroviral (ARV)-experienced HIV-infected patients successfully treated with raltegravir (RAL)-containing therapy. Nineteen patients with virological failure whose ARV treatment was switched to a RAL-based salvage regimen with virological success were included (Group I). Ten patients in virological failure and responding to ARV salvage therapy not containing RAL were also included (Group II). The HIV-1 DNA level in peripheral blood mononuclear cells (PBMC) was assessed by real-time PCR at baseline, W12, W24, W36 or W48. In group I, a marked decrease in the HIV-1 DNA level was observed at W12 both in PBMC (median decrease = 0.38 log(10)copies/10(6)PBMC; P < 0.001) and in CD4 T cells (0.85 log(10)copies/10(6)CD4 T cells; P < 0.001). Plasma HIV-1 RNA decrease was correlated with HIV-1 DNA decrease expressed as copies/10(6)CD4 T cells (r = 0.55, P = 0.03). HIV-1 DNA level reached a steady state by W24. Thus, RAL-containing treatment in highly ARV-experienced patients was associated with a rapid HIV-1 DNA decrease, mainly in the circulating CD4 T cells compartment. Group II patients showed an early decrease in the HIV-1 DNA load until W12, which was 2.5-fold less pronounced in the CD4 T cells compartment than in the RAL-treated patients. The potent action of RAL-containing treatment observed in the CD4 T cells compartment may suggest a pronounced reduced inhibition in the pool of regenerating CD4 T cells on a RAL-based therapy.

Drug resistance profiles for the HIV integrase gene in patients failing raltegravir salvage therapy.

OBJECTIVES: Recent data showed the selection of mutations in the integrase gene, mainly involving position 148 or 155, in patients displaying virological failure (VF) on raltegravir (RAL) therapy. Here, we describe the development of RAL resistance, in both plasmatic and cellular compartments, in three heavily pretreated HIV-infected patients failing RAL-containing regimens. METHODS: Three of 17 patients receiving RAL displayed VF. The entire integrase gene, isolated from plasma and peripheral blood mononuclear cells (PBMC), was sequenced. A clonal analysis was performed in one patient at the time of VF. RESULTS: At the time of VF, RAL-resistance mutations were selected: (i) Q148R in patients 1 and 3; (ii) T66A and E92Q in patient 2. A gradual accumulation of new mutations was observed in all patients, including G140S, Q148H and N155H in patient 1, L74I in patient 2, and G140S in patient 3. Clonal analysis showed the coexistence, in patient 1, of the two common resistance pathways (mutations Q148R/H and N155H) found in distinct quasi-species. In addition, RAL-resistance mutations were detected in HIV DNA in all patients. CONCLUSIONS: Having rapidly established, resistance to RAL evolves and diversifies, and is likely to impact the efficacy of subsequently used second-generation integrase inhibitors. Moreover, RAL-resistance mutations can be archived early in PBMC.

Cervicovaginal secretory antibodies to human immunodeficiency virus type 1 (HIV-1) that block viral transcytosis through tight epithelial barriers in highly exposed HIV-1-seronegative African women.

Antibodies to human immunodeficiency virus (HIV) of the IgA, IgG, and IgM isotypes and high levels of the HIV suppressive beta-chemokine RANTES (regulated on activation, normally T cell expressed and secreted) were found in the cervicovaginal secretions (CVSs) of 7.5% of 342 multiply and repeatedly exposed African HIV-seronegative female sex workers. The antibodies are part of a local compartmentalized secretory immune response to HIV, since they are present in vaginal fluids that are free of contaminating semen. Cervicovaginal antibodies showed a reproducible pattern of reactivity restricted to gp160 and p24. Locally produced anti-env antibodies exhibit reactivity toward the neutralizing ELDKWA epitope of gp41. Study results show that antibodies purified from CVSs block the transcytosis of cell-associated HIV through a tight epithelial monolayer in vitro. These findings suggest that genital resistance to HIV may involve HIV-specific cervicovaginal antibody responses in a minority of highly exposed HIV-seronegative women in association with other protecting factors, such as local production of HIV-suppressive chemokines.

Quantitation of human immunodeficiency virus type 1 (HIV-1) RNA in cell-free cervicovaginal secretions: comparison of reverse transcription-PCR amplification (AMPLICOR HIV-A MONITOR 1.5) with enhanced-sensitivity branched-DNA assay (Quantiplex 3.0).

Two commercially available hypersensitive assays for human immunodeficiency virus type 1 (HIV-1) RNA quantitation, AMPLICOR HIV-1 Monitor Test 1.5 and Quantiplex HIV RNA 3.0, were compared to detect and quantify HIV-1 RNA in the cell-free fraction of cervicovaginal secretions collected by vaginal washing. Three panel specimens were used: pooled cervicovaginal secretions spiked with HIV-1 subtype A or HIV-1 subtype B and cervicovaginal lavages from HIV-positive and HIV-negative women. Compared to the AMPLICOR HIV-1 Monitor Test 1.5 assay, the Quantiplex HIV-1 3.0 assay yielded higher estimates of HIV-1 RNA concentrations in several tested samples spiked with HIV-1 RNA subtype A, as well as subtype B, particularly samples containing low amounts of HIV-1 RNA. The sensitivity and specificity of the AMPLICOR HIV-1 Monitor Test 1.5 assay were 93 and 100%, respectively; the sensitivity and specificity of the Quantiplex HIV RNA 3.0 assay were 97 and 50%, respectively. In conclusion, in quantifying HIV-1 RNA in cervicovaginal secretions, the Quantiplex HIV RNA 3.0 may lack specificity, and the AMPLICOR HIV-1 Monitor Test 1.5 assay, although highly specific, may lack sensitivity.

In vitro inactivation of Chlamydia trachomatis and of a panel of DNA (HSV-2, CMV, adenovirus, BK virus) and RNA (RSV, enterovirus) viruses by the spermicide benzalkonium chloride.

Kinetics of inactivation by the detergent spermicide benzalkonium chloride (BZK) of Chlamydia trachomatis and of a panel of DNA viruses [herpes simplex virus hominis type 2 (HSV-2), cytomegalovirus (CMV), adenovirus (ADV) and BK virus (BKV)] and RNA [respiratory syncytial virus (RSV) and enterovirus (ENV)] were established in accordance with a standardized in vitro protocol. After a 5 min incubation, inactivation of >95% of HSV-2 and CMV was obtained at a concentration of 0.0025% (w/v) (25 Ig/L); concentrations as low as 0.0005%, 0.0050% and 0.0125%, induced a 3.0 log10 reduction in infectivity of HSV-2 and CMV, RSV and ADV, respectively. After a 60 min incubation, concentrations of 0.0125% and 0.050% provided a 3.0 log10 reduction in infectivity of ENV and BKV, respectively. These features indicate that sensitivity to BZK was very high (HSV-2 and CMV) or high (RSV) for enveloped viruses, intermediate (ADV) or low (ENV and BKV) for non-enveloped viruses. Furthermore, BZK had marked antichlamydial activity, showing >99% killing after only a 1 min incubation at a concentration of 0.00125%. BZK demonstrates potent in vitro activity against the majority of microorganisms causing sexually transmitted infectious diseases, including those acting as major genital cofactors of human immunodeficiency virus transmission. These attributes qualify BZK as a particularly attractive candidate for microbicide development.

The cumulative occurrence of resistance mutations in the HIV-1 protease gene is associated with failure of salvage therapy with ritonavir and saquinavir in protease inhibitor-experienced patients.

Salvage therapy with ritonavir (RTV) and saquinavir (SQV) failed to achieve virological and immunological improvement in 24 HIV-infected patients who discontinued triple therapy with RTV or indinavir (IDV) because of failure or intolerance to treatment. Changes in the HIV-1 protease gene sequence were analyzed prospectively in 14 patients. No primary protease mutation was found prior to the use of protease inhibitors. After 7 months of treatment with IDV or RTV, primary resistance mutations at codons pol 46 and/or pol 82 were observed in 11 of 13 patients. After 16 weeks on RTV-SQV, novel primary mutations related to SQV emerged in 7 of 13 patients, together with an increase in the number of secondary resistance mutations. Our observations indicate that the cumulative occurrence of resistance mutations in the protease gene was associated with failure of antiretroviral therapy. The presence of mutations to a first protease inhibitor may represent a risk factor for the failure of a subsequent treatment with a second line protease inhibitor.

Selection of drug-resistant variants in the female genital tract of human immunodeficiency virus type 1-infected women receiving antiretroviral therapy.

We investigated human immunodeficiency virus (HIV) type 1 RNA, proviral DNA, and antiretroviral drug-resistant variants in cervicovaginal secretions of HIV-1-infected women receiving antiretroviral therapy. The prevalence of detectable HIV-1 RNA in genital secretions was inversely related to the number of antiretroviral drugs taken by the patients. Proviral DNA was detected in approximately half of all samples of cervicovaginal secretions from HIV-1-infected women, regardless of the presence or absence of HIV-1 RNA in cervicovaginal secretions and of the antiretroviral regimen. In cervicovaginal secretions of most women with persisting genital viral replication, HIV variants exhibiting mutations associated with drug resistance against protease and reverse-transcriptase pol genes were found. Our observations indicate that antiretroviral therapy is not effective in purging the female genital tract of cell-associated provirus and that antiretroviral drugs that penetrate the female genital tract at suboptimal concentrations exert a potent selective pressure on genital HIV variants when local replication of free HIV-1 RNA persists.

High levels of drug-resistant human immunodeficiency virus variants in patients exhibiting increasing CD4+ T cell counts despite virologic failure of protease inhibitor-containing antiretroviral combination therapy.

The genotypic mutations associated with indinavir resistance were analyzed in 27 patients who exhibited sustained CD4+ T cell responses to highly active antiretroviral therapy (HAART), despite virologic failure of treatment. After 12 months of HAART, 1 or 2 primary resistance mutations had occurred in 18 (66%) of the patients, and secondary mutations had accumulated in 22 (88%) of the patients. The number and patterns of mutations in the patients who exhibited discrepant responses to HAART did not differ from those observed in patients who exhibited immunologic and virologic failure to therapy. Results indicate that many patients have prolonged immunologic benefits, despite the development of virologic failure and protease inhibitor mutations. The clinical course of this group of patients calls into question the relevance of genotypic resistance and plasma human immunodeficiency virus RNA level as surrogate markers in patients receiving HAART.

Efficacy of a five-drug combination including ritonavir, saquinavir and efavirenz in patients who failed on a conventional triple-drug regimen: phenotypic resistance to protease inhibitors predicts outcome of therapy.

OBJECTIVE: to assess the safety and efficacy of a combination of ritonavir, efavirenz and two recycled nucleosides in patients who failed on a conventional triple-drug regimen including indinavir or ritonavir. METHODS: An open label study of ritonavir (100 mg twice daily), saquinavir (1000 mg twice daily), efavirenz (600 mg per day) and nucleoside analogues in 32 saquinavir- and efavirenz-naive protease inhibitor-experienced patients. Patients were included on the basis of plasma levels of HIV RNA above 5000 copies/ml while on conventional antiretroviral therapy. Phenotypic resistance and genotypic resistance mutations to saquinavir were assessed at baseline. Peak and trough plasma levels of saquinavir were monitored throughout the study. RESULTS: Median CD4 cell counts and median plasma HIV RNA at baseline were 258 x 10(6)/l and 4.31 log10 copies/ml, respectively. The plasma viral load decreased by a median of 1.20 log10 copies/ml and the CD4 cell count increased by a median 60 x 10(6) cells/l at week 24 of therapy. Seventy-one per cent of the patients achieved a plasma viral load < 500 copies/ml and 45% achieved a viral load < 50 copies/ml. Patients exhibiting phenotypic resistance to saquinavir at baseline experienced a median decrease in HIV RNA of 0.91 log10 copies/ml at week 24 of therapy, as compared with a decrease of 1.52 log10 copies/ml in those exhibiting sensitive viral strains (P = 0.03). Genotypic resistance to saquinavir was not predictive of virologic failure. CONCLUSION: Our results indicate that the combination of ritonavir, saquinavir and efavirenz is safe and effective at 24 weeks in over two-thirds of patients who previously failed on highly active antiretroviral therapy, and that the determination of phenotypic resistance may be of greater value than the detection of resistance mutations to predict the outcome of salvage therapy in this setting.

The sequential occurrence of pol 215 and pol 41 zidovudine resistance mutations is associated in an additive fashion with low CD4 cell counts and high plasma and cellular HIV viral load.

We report on a cross-sectional study of virological and immunological surrogate markers of HIV infection in 115 patients for whom a determination of the pol 215 and pol 41 zidovudine (ZDV) resistance mutations had been described between January 1995 and February 1996. The patients received ZDV alone or a combination of ZDV and zalcitabine or didanosine. A total of 55, 15 and 45 patients exhibited a wild (W), a mixed (MIX) or a mutant (M) genotype at codon pol 215, respectively; 85, 10 and 20 patients exhibited a W, a MIX or a M genotype at codon pol 41, respectively. Patients exhibiting the pol 215 M genotype had lower CD4 cells, higher plasma viral load and higher proviral burden than patients exhibiting the pol 215 W genotype. Patients who had variants exhibiting both pol 215 M and pol 41 M or MIX genotypes had significantly worsened surrogate marker values than patients having variants only carrying the pol 215 M genotype. These observations demonstrate that the two mutations additively associate with pejorative surrogate markers.

High prevalence in Central Africa of blood donors who are potentially infectious for human herpesvirus 8.

BACKGROUND: In western countries, the transmission of human herpesvirus 8 (HHV-8) via blood transfusion has been recently postulated. In sub-Saharan African, the incidence of HHV-8-associated Kaposi's sarcoma and the seroprevalence for HHV-8 in autochthonous populations are high. STUDY DESIGN AND METHODS: The aim of this study was to estimate the prevalence of blood donations potentially infectious for HHV-8 in the general adult population of Central Africa. Forty-nine blood donors at the Centre de Transfusion Sanguine in Bangui, the capital of the Central African Republic, were selected. Forty-five inpatients of Broussais Hospital, Paris, France, who were known to be seronegative for HIV and hepatitis B and C viruses and who had not received heart or kidney transplants, were chosen as a European "control" group for comparison. HHV-8 DNA sequences were detected in peripheral blood mononuclear cells by nested polymerase chain reaction using primer sets located in the HHV-8 open reading frame 26. RESULTS: Eleven (22.5%; 95% CI: 12%-37%) of 49 blood donors were positive for HHV-8. Three (6%) were HIV-1 seropositive. Two (67%) of the 3 HIV-infected blood donors were also positive for HHV-8. All blood donors were apparently healthy; none was known to suffer from Kaposi's sarcoma. Only one (2.2%) control was carrying HHV-8 DNA on the peripheral blood mononuclear cells. The prevalence of HHV-8 was higher in blood donors from Bangui than in patients from Broussais Hospital. CONCLUSIONS: HHV-8 infection is highly prevalent in an apparently healthy adult population from Central Africa, which raises concerns about HHV-8 transmission through transfusion in this setting.

Genetically related human immunodeficiency virus type 1 in three adults of a family with no identified risk factor for intrafamilial transmission.

A small number of cases of human immunodeficiency virus (HIV) infection have been reported in individuals with no identified risk factors for transmission. We report on the seroconversion of the 61-year-old mother and the subsequent finding of HIV seropositivity in the 66-year-old father of a 31-year-old AIDS patient. Extensive investigation failed to identify any risk factor for intrafamilial transmission. We conducted a genetic analysis and determined the amino acid signature patterns of the V3, V4, and V5 hypervariable domains and flanking regions in the HIV-1 gp120 env gene of 26 clones derived from proviral DNA in peripheral blood mononuclear cells of the members of the family. env sequences of the viruses isolated from the patients were compared with sequences of HIV-1 subtype B viruses from Europe and local field isolates. Phylogenetic analysis revealed that the sequences of the viruses isolated from the patients were genetically related and formed an intrafamilial cluster of HIV-1 distinct from other subtype B viruses. Interindividual nucleotide variability in the C2-V3 and V4-C4-V5 domains ranged between 1.2 and 5.0% and between 2.2 and 7.5%, respectively, whereas divergence between HIV strains from the patients and control viral strains ranged from 6.6 to 29.3%. The amino acid signature patterns of viral clones from the three patients were closely related. In the C2-V3 region, two minor clones derived from the son's virus showed less nucleotide divergence (mean, 3.5 and 3.9%) than did the clones derived from the viruses of both parents or the seven other predominant clones derived from the virus from the son (mean, 5.4%). The top of the V3 loop of the last two clones and of all viral clones from the parents exhibited an unusual GPGG sequence. This is the first report of genotypic relatedness of HIV-1 in three adults of the same family in the absence of identified risk factor for transmission between the members of the family. Our findings suggest that atypical transmission of HIV may occur.

Discrepant responses to triple combination antiretroviral therapy in advanced HIV disease.

OBJECTIVES: To determine the clinical, virological and immunological outcome in a cohort of unselected patients receiving triple combination therapy for more than 1 year. METHODS: Prospective follow-up of a cohort of 162 unselected, protease inhibitor-naive, antiretroviral-experienced patients with advanced HIV disease, treated with indinavir combined with two nucleoside analogues. RESULTS: The mean CD4 cell count and plasma HIV RNA level in the study group at baseline were 69+/-5 x 10(6)/l and 4.75+/-0.07 log10 copies/ml, respectively. Five per cent of patients died prematurely or were lost to follow-up. Fifty-seven per cent of patients responded to therapy, as assessed by a sustained increase in CD4 cell counts above 50 x 10(6)/l and a decrease in plasma HIV RNA greater than 1 log10 copies/ml, throughout 12.1 months of follow-up. Seventeen per cent of patients were immunological and virological non-responders. Twenty-one per cent of patients exhibited discrepant virological and immunological responses to treatment, of whom one-half failed to exhibit significant increases in CD4 cells despite a virological response to therapy and one-half exhibited increased CD4 cell counts in the absence of significant decrease in plasma viral load. The incidence of AIDS-defining events in the latter group of patients was similar to that of responder patients, whereas their incidence was higher in patients who failed to exhibit a virological and immunological response and those who failed to increase CD4 cells despite a significant decrease in viral load. CONCLUSION: Our observations of discrepant immunological and virological responses to treatment raise the issue of the significance of persistent elevated levels of plasma HIV RNA and of the relevance of measurements of plasma viral load for assessing the efficacy of antiretroviral therapy in patients whose CD4 cell counts increase despite the absence of significant decrease in plasma HIV viral load.