Luiz Claudio Arraes

Therapeutic dendritic-cell vaccine for chronic HIV-1 infection

We present the results of a preliminary investigation of the efficacy of a therapeutic dendritic cell (DC)-based vaccine for HIV-1. We immunized 18 chronically HIV-1-infected and currently untreated individuals showing stable viral loads for at least 6 months with autologous monocyte-derived DCs loaded with autologous aldrithiol-2-inactivated HIV-1. Plasma viral load levels were decreased by 80% (median) over the first 112 d following immunization. Prolonged suppression of viral load of more than 90% was seen in 8 individuals for at least 1 year. The suppression of viral load was positively correlated with HIV-1-specifc interleukin-2 or interferon-γ-expressing CD4+ T cells and with HIV-1 gag–specific perforin-expressing CD8+ effector cells, suggesting that a robust virus-specific CD4+ T-helper type 1 (TH1) response is required for inducing and maintaining virus-specific CD8+ effectors to contain HIV-1 in vivo. The results suggest that inactivated whole virus–pulsed DC vaccines could be a promising strategy for treating people with chronic HIV-1 infection.

DEFB1 gene polymorphisms and increased risk of HIV-1 infection in Brazilian children

In our study we analysed three single nucleotide polymorphisms (SNPs) in the 5′ untranslated region (UTR) of the DEFB1 gene, namely −52(G/A) −44(C/G) and −20(G/A), in three groups of northeastern Brazilian children in order to assess their role in HIV-1 infection. Our results allowed us to hypothesize that the SNPs located in the 5′ UTR of the DEFB1 gene can be employed as a marker of risk for HIV-1 infection.

Association between HLA-G 3'UTR 14-bp polymorphism and HIV vertical transmission in Brazilian children.

Objectives:The aim of our study was to verify the possible association between an HLA-G 14-bp deletion/insertion polymorphism and perinatal HIV transmission in Brazilian children. Design:We analyzed the 14-bp deletion/insertion polymorphisms in seronegative (i.e., exposed uninfected, N = 71) and seropositive (exposed infected, N = 175) Brazilian children born from HIV-positive mothers and in healthy controls (n = 175). Methods:HLA-G 14-bp deletion/insertion polymorphism (rs16375) was detected by PCR amplification of the target sequence followed by agarose gel electrophoresis. All the samples were also analyzed by direct sequencing in order to validate the genotyping results. Results:HIV-exposed uninfected children showed significant differences in their allele and genotype frequencies of the HLA-G 14-bp polymorphism when compared to both seropositive children and healthy controls. The 14-bp-deleted (D) allele was more frequent in exposed uninfected children (79%) than in healthy controls (60%) and HIV-positive children (58%); the higher percentage of the D allele found in the exposed uninfected children with respect to HIV-positive individuals was significantly associated with a reduced risk of vertical transmission. This effect was ascribable to the presence of the D/D homozygous genotype. Conclusion:Our findings support the possible role for the HLA-G 14-bp deletion/insertion polymorphism in the HIV vertical transmission in Brazilian children. The presence of the D allele and D/D genotype is associated with a protective effect toward HIV perinatal infection.

The biological meaning of anti-HBC positive result in blood donors: relation to HBV-DNA and to other serological markers

In order to assess the potential risk of anti-HBc-positive blood donors for post-transfusional hepatitis and to investigate whether other HBV serological markers are capable of identifying the presence of the virus, 1000 first-time blood donors were enrolled between June and July 1997. These donors were screened using routine Brazilian blood center tests (HIV 1 and 2, HTLV 1 and 2, Chagas disease, Syphilis, HCV, HBsAg, anti-HBc and ALT ). The 120 (12%) found to be anti-HBc-positive underwent further tests: HBe, anti-HBe, anti-HBs and HBV-DNA by PCR. Ten cases were HBsAg positive and all were HBV-DNA positive by PCR. Three HBsAg-negative donors were HBV-DNA-positive. Two HBV-DNA-positive donors were also anti-HBs-positive. All the HBV-positive donors had at least one HBV marker other than anti-HBc. Anti-HBc is an important cause of blood rejection. Testing for HBsAg alone is not fully protective and anti-HBc remains necessary as a screening test. The presence of anti-HBs is not always indicative of absence of the virus. The addition of other HBV serological markers could represent an alternative in predicting the presence of the virus when compared with PCR. It is recommended that other studies should be carried out to confirm this finding.

IL-18 gene promoter polymorphism is involved in HIV-1 infection in a Brazilian pediatric population

In our study, we identified a polymorphism (C-607A) in the promoter region of the IL-18 gene that shows different frequencies between human immunodeficiency virus (HIV)-1-infected children and healthy controls in a pediatric Brazilian population. The presence of the −607 C allele correlates to HIV-1 infection and confers an increased risk of infection in subjects carrying the single nucleotide polymorphism.

A cost-effective melting temperature assay for the detection of single-nucleotide polymorphism in the MBL2 gene of HIV-1-infected children

We report a fast (less than 3 h) and cost-effective melting temperature assay method for the detection of single-nucleotide polymorphisms in the MBL2 gene. The protocol, which is based on the Corbett Rotor Gene real time PCR platform and SYBR Green I chemistry, yielded, in the cohorts studied, sensitive (100%) and specific (100%) PCR amplification without the use of costly fluorophore-labeled probes or post-PCR manipulation. At the end of the PCR, the dissociation protocol included a slow heating from 60 degrees to 95 degrees C in 0.2 degrees C steps, with an 8-s interval between steps. Melting curve profiles were obtained using the dissociation software of the Rotor Gene-3000 apparatus. Samples were analyzed in duplicate and in different PCR runs to test the reproducibility of this technique. No supplementary data handling is required to determine the MBL2 genotype. MBL2 genotyping performed on a cohort of 164 HIV-1-positive Brazilian children and 150 healthy controls, matched for age and sex and ethnic origin, yielded reproducible results confirmed by direct sequencing of the amplicon performed in blind. The three MBL2 variants (Arg52Cys, Gly54Asp, Gly57Glu) were grouped together and called allele 0, while the combination of three wild-type alleles was called allele A. The frequency of the A/A homozygotes was significantly higher among healthy controls (0.68) than in HIV-infected children (0.55; P = 0.0234) and the frequency of MBL2 0/0 homozygotes was higher among HIV-1-infected children than healthy controls (P = 0.0296). The 0 allele was significantly more frequent among the 164 HIV-1-infected children (0.29) than among the 150 healthy controls (0.18; P = 0.0032). Our data confirm the association between the presence of the mutated MBL2 allele (allele 0) and HIV-1 infection in perinatally exposed children. Our results are in agreement with the literature data which indicate that the presence of the allele 0 confers a relative risk of 1.37 for HIV-1 infection through vertical transmission.

HPV31 and HPV33 incidence in cervical samples from women in Recife, Brazil.

Human papillomavirus (HPV) has been extensively studied concerning genomic structure, infection mechanisms, and diversity of types, as well as disease progression stages and development of vaccines. HPV type prevalence can differ in specific populations in different countries, according to ethnicity. This is the first report of an integrated project to evaluate the incidence of HPV types in different regions in Brazil in order to obtain data for vaccine development. Cervical samples were collected from women seen at a public hospital in Pernambuco, Northeast Brazil, for routine evaluation of genital alterations. Selection of the patients was random. There was a strong prevalence of HPV16 and a high incidence of HPV types 31 and 33. These data foster the discussion about the need to evaluate viral prevalence in each geographic region in order to develop targeted vaccine programs.

Association between MBL2 gene functional polymorphisms and high-risk human papillomavirus infection in Brazilian women

We studied the association between high-risk human papillomavirus (HPV) infection and MBL2 functional polymorphisms in a group of 180 high-risk HPV-infected women and 180 healthy control subjects. The most frequent high-risk HPV genotypes were 16 (47.2%), 31 (11.7%), 33 (5%), and 18 (2.2%), respectively. Of the 180 HPV-infected women, 99 presented with uterine cervical cancer and 81 did not. No differences in MBL2 genotype or in allelic or haplotype frequencies were found between HPV patients who developed cervical uterine cancer and those who did not. When considering combined genotypes grouped according to MBL production (designated as high, low, and deficient producers), we detected a significant difference between healthy controls and high-risk HPV-positive patients, the latter group showing increased frequencies of deficient-producer genotypes (14.4% vs 9.4% in the healthy control group, corrected p = 0.04). In conclusion, a correlation between MBL2 polymorphisms and high-risk HPV infection was found in this study.

Frequency of HLA B*5701 allele carriers in abacavir treated-HIV infected patients and controls from northeastern Brazil

In clinical practice, using pharmacogenetics has risen in importance since the technologies for genomic variation searches for different responses to drugs became widespread, easier, more rapid, and affordable for a specialized laboratory. The main pharmacogenetics studies have mostly involved drug resistance monitoring in oncology patients;1 however, other applications are being developed to detect genetic molecular markers associated with resistance or hypersensitivity/adverse reactions to antiretroviral drug treatment in Human Immunodeficiency Virus (HIV)-infected patients.2 HIV treatment is known to be limited by adverse drug reactions and the development of resistance. One significant example is the strong association of the Abacavir hypersensitivity reaction with HLA-B*5701 in HIV-positive patients.3 Abacavir, a common drug for treating HIV-infected patients, is an efficient nucleoside reverse-transcriptase inhibitor with a beneficial long-term toxicity profile, often used with other antiretroviral agents. However, Abacavir can yield adverse effects such as immunologically-driven hypersensitivity in 5 to 8% of HIV-positive subjects during the first six weeks of use.2 Hypersensitivity symptoms immediately reverse after the interruption of Abacavir.4 The hypersensitivity reaction to Abacavir has been reportedly associated with the presence of the major histocompatibility complex class I allele HLA-B*5701 and this association has been confirmed in several replication studies in different ethnic groups of HIV-positive patients.3 In our study, we searched for the presence of HLA B*5701 in 96 HIV-positive patients treated with Abacavir and in 243 healthy subjects from Northeastern Brazil (Recife, Pernambuco) to verify the percentage of HLA B*5701 allele carriers in HIV patients and in the general population from Northeast Brazil. This area is known to harbor a tri-hybrid population resulting from contributing African (44%), Caucasian (34%), and native American (22%) genomes.5

Polymorphisms in innate immunity genes and patients response to dendritic cell-based HIV immuno-treatment

Dendritic cells (DCs)-based vaccine was demonstrated to increase HIV specific cellular immune response; however, in some HIV-infected patients, the response to the vaccine resulted to be not effective. In order to understand if the outcome of the vaccination may be influenced by the hosts genome and natural immunity, we studied the innate immune genome of HIV-infected patients previously vaccinated with DCs. We identified 15 SNPs potentially associated with the response to the immuno-treatment and two SNPs significantly associated with the modulation of the response to the DC vaccine: MBL2 rs10824792 and NOS1 rs693534. These two SNPs were also studied in different ethnic groups (Brazilians, African and Caucasian) of HIV-infected, exposed uninfected and unexposed uninfected subjects. The HIV positive Caucasian patients were also characterized by different disease progressions. Our findings suggest that, independently and/or in addition to other variables, the hosts genome could significantly contribute to the modulation of the response to the DC vaccine.