A Baseline Analysis of Regulatory Review Timelines for ANVISA: 2013-2016.

BACKGROUND: The Brazilian health regulatory agency (Agência Nacional de Vigilância Sanitária, ANVISA) has embarked on transformational initiatives to fulfill its mandate to provide timely access to safe, effective, and quality therapeutics. A new Brazilian law was enacted to provide the agency with greater flexibility. Optimizing Efficiencies in Regulatory Agencies (OpERA) is a regulatory-strengthening program that seeks to provide benchmarking data that can be used to define performance targets and focus performance improvement. The objective of this study was to use OpERA methodology to undertake a retrospective analysis of the timelines associated with important components of the ANVISA regulatory review process to establish a baseline against which the influence of the new law could be measured. METHODS: The OpERA tool was used to collect specific milestone data that identify time periods, review stages, and data points for products approved by ANVISA 2013-2016. RESULTS: For the 138 products approved in this cohort, the overall median approval time was 795 days. ANVISA and submitting companies will need to reduce their review and response times by approximately half in order to meet the total time goal of 365 days. CONCLUSIONS: The observations from this baseline study have identified opportunities for ANVISA and sponsor companies to collaborate to reduce regulatory assessment times while assuring the timely approval of safe and effective, quality medicines. These analyses will be repeated to determine how the provisions of the new Law will impact the activities of ANVISA and the extent of sponsors' contributions to this effort.

Phylogeny and polymorphism in the long control region, E6, and L1 of human papillomavirus types 53, 56, and 66 in central Brazil.

INTRODUCTION: Several studies related that different human papillomavirus (HPV) types and intratype variants can present different oncogenic potential. In opposite to HPVs 16 and 18 variants, information about variants of other carcinogenic HPV types is still scarce. The aim of this study was to investigate the genetic variability of HPVs 53, 56, and 66 from Central Brazil isolates. METHODS: The long control region (LCR), E6, and L1 genomic regions were amplified and sequenced. We evaluate for nucleotide variations in relation to the reference sequence of each HPV type and also the conservation of physicochemical properties of the deduced amino acid substitutions. In silico analysis was performed to locate binding sites for transcriptional factors within the LCR. Moreover, we performed a phylogenetic analysis with the Central Brazilian and worldwide sequences available at genomic databases. RESULTS: Gathering LCR, E6, and L1 genomic regions, the highest genetic variability was found among HPV-53 isolates with 52 nucleotide variations, followed by HPVs 56 and 66 with 24 and 16 nucleotide substitutions, respectively. The genetic analysis revealed 11 new molecular variants of all HPV types analyzed, totalizing 31 new nucleotide and 3 new amino acid variations. Eight nonconservative amino acid substitutions were detected, which may indicate a biological and pathogenic diversity among HPV types. Furthermore, 8 nucleotide substitutions were localized at putative binding sites for transcription factors in the LCR with a potential implication on viral oncogene expression. The HPVs 53, 56, and 66 phylogenetic analysis confirmed a dichotomic division only described to HPV subtypes and different from the patterns described for HPVs 16 and 18 variants. CONCLUSIONS: The high genetic variability observed emphasizes the importance of investigating polymorphisms in types other than HPVs 16 or 18 to better understand the molecular genomic profile of viral infection by different HPV types.

Genetic variability and phylogeny of the high-risk HPV-31, -33, -35, -52, and -58 in central Brazil.

More than 100 HPV types have been described, 13 of which are classified as high-risk due to their association with the development of cervical cancer. The intratype genomic diversity of HPV-16 and -18 has been studied extensively, while little data have been generated for other less common high-risk types. The present study explores the nucleotide variability and phylogeny of the high-risk HPV-31, -33, -35, -52, and -58, in samples from Central Brazil. For this purpose, the LCR and the E6 and L1 genes were sequenced. Several variants of these HPV types were detected, some of which have been detected in other parts of the world. Furthermore, new variants of all types examined were characterized in a total of 13 new variants. Based on the E6 and L1 sequences, variants were described comprising conservative and non-conservative amino acid changes. For phylogenetic tree construction, samples characterized in this study were compared to others described and submitted to GenBank previously. The phylogenetic analysis of HPV-31, -33, -35, and -58 isolates did not reveal ethnic or geographical clustering as observed previously for HPV-16 and -18. HPV-35 analysis showed a dichotomic branching characteristic of viral subtypes. Interestingly, four clusters relative to the analysis of HPV-52 isolates were identified, two of which could be classified as Asian and European branches. The genomic characterization of HPV variants is crucial for understanding the intrinsic geographical relatedness and biological differences of these viruses and contributes further to studies on their infectivity and pathogenicity.

New variants of human papillomavirus type 18 identified in central Brazil.

HPV-18 is the second most prevalent human papillomavirus genotype found in cervical cancer. Nucleotide variations in HPV-18 sequence can interfere with the viral oncogenic potential. However, the knowledge about HPV-18 variants in Brazil is still limited. The present study aims to determine the LCR, E6, and L1 genetic variability of HPV-18 variants found in women co-infected with HIV-1 in Central Brazil. Four HPV-18 samples were identified and had the LCR, E6, and L1 genomic regions sequenced. It was possible to characterize three European variants and one African variant of HPV-18. All of them are new variants, showing nucleotide substitutions not previously reported. Nucleotide variations in binding sites for transcriptional factors were observed. Phylogenetic analysis was also performed, evidencing the three clusters related to the Asian-American, African, and European variants. The characterization of HPV genetic variability is of pivotal importance to the understanding of the viral pathogenicity.

Caracterização molecular do papilomavírus humano em mulheres infectadas com o vírus da imunodeficiência humana do tipo 1 no Distrito Federal e Entorno / Molecular characterization of human papillomavirus in women infected with hiv-1 in Central Brazil

Introdução: a infecção por HPV é a infecção transmitida sexualmente mais comum no mundo e representa uma das infecções oportunistas que mais acomete as mulheres portadoras do HIV-1. A diversidade genética do HPV foi investigada entre mulheres infectadas pelo HIV-1 no Distrito Federal e Entorno. Método: duzentas amostras cervicais de mulheres soropositivas para o HIV-1 foram submetidas a exame citopatológico e à amplificação por PCR. A genotipagem do HPV foi realizada por RFLP e as análises estatísticas pelo programa Epi Info. Resultados: a prevalência do HPV foi 41 por cento. Entre as amostras sem anormalidades citopatológicas cervicais, 29,2 por cento tinham HPV, enquanto a maioria das amostras com citologia alterada tinha HPV (89,7 por cento). Uma grande diversidade de genótipos foi observada com a maior prevalência dos HPV -16 e -81 (12,2 por cento). Observou-se uma prevalência significativamente maior de HPV em mulheres com menos de 30 anos. As análises estatísticas sugeriram que a imunossupressão estava associada com a prevalência de anormalidades citológicas, mas não com a presença de infecção por HPV. A carga viral do HIV-1, por sua vez, estava associada com a detecção de DNA do HPV e de anormalidades citológicas. Não foi observada associação significativa entre a terapia anti-retroviral e a presença de HPV ou de anormalidades citológicas. Conclusão: uma proporção elevada de mulheres soropositivas para oHIV-1 está co-infectada por HPV, e, muitas vezes, por genótipos oncogênicos, mesmo em casos onde a avaliação citológica não revela resultados anormais, demonstrando a necessidade de acompanhamento dessa população. Palavras-chave: HPV; HIV; Distrito Federal.

Resistência do vírus da imunodeficiência humana tipo 1 aos inibidores de protease em amostragem do Laboratório Central de Saúde Pública do Distrito Federal no ano de 2002 / Resistance of the human immunodeficincy virus type 1 to protease inhibitors in samples from Laboratório Central de Saúde Pública do Distrito Federal from 2002

A pandemia da AIDS é um dos maiores problemas de saúdepública no mundo e a resistência aos anti-retrovirais, um dos principais obstáculos para o tratamento efetivo da infecção pelo vírus da imunodeficiência humana tipo 1 (HIV-1). Os estudos sobre a análise da resistência a esses medicamentos são particularmente importantes parao manejo clínico de indivíduos infectados pelo HIV-1. O HIV codifica uma protease que é essencial para a maturação das partículas virais e, por isso, essa proteína é alvo de abordagens terapêuticas utilizandoseos inibidores de protease. Entretanto, muitas mutações no gene da protease do HIV-1 têm sido associadas à resistência aos anti-retrovirais,conduzindo à falhas do tratamento. Objetivo: Descrever as mutações no gene da protease e os perfis de resistência genotípica em indivíduos infectados pelo HIV-1 provenientes do Distrito Federal e Entorno. Método: O gene da protease de 28 isolados de indivíduos infectados por HIV-1 foi amplificado por nested PCR e seqüenciado. As seqüênciasforam analisadas utilizando-se o programa HIVdb para determinação da susceptibilidade às drogas. Resultados: A mutação primária I50V foi a mais prevalente, ocorrendoem 28.57 por cento das amostras. Esta mutação está associada ao medicamento Amprenavir, que apresentou um alto nível de resistência.Conclusão: Esses resultados estão de acordo com uma descrição previamente realizada sobre a resistência aos inibidores de protease no Distrito Federal, e devem ser aprofundados para auxiliar no tratamento de indivíduos infectados pelo HIV-1. Palavras-chave: AIDS, HIV-1, inibidores de proteases.

New HPV-16 European and non-European variants in Central Brazil.

HPV-16 is the most prevalent human papillomavirus genotype found in cervical intraepithelial neoplasias. The regulatory region of the HPV genome, LCR, has several binding sites for cellular and viral transcription factors, and nucleotide substitutions in this genomic region can interfere with the viral oncogenic expression. The present study aims to determine the LCR variability of European and non-European HPV-16 variants found in Brazil. Through automated sequencing, it was possible to characterize the LCR of ten non-European (eight Asian-American, one African 1, one African 2) and twelve European isolates. Among the 22 isolates analyzed, nine may be new variants of HPV-16, with different combinations of previously reported nucleotide substitutions, and three showed new substitutions not previously reported. Two new nucleotide substitutions, the insertion of T at position 7621 and the substitution of A to G at position 7836, were found in a single isolate, Bsb-14, a putative new African 1 variant. The characterization of the LCR of human papillomaviruses can be of pivotal importance to the understanding of the viral replication and pathogenicity.

HIV-1 subtypes and mutations associated to antiretroviral drug resistance in human isolates from Central Brazil

A detecção de polimorfismos do HIV-1 que estejam associados à resistência às drogas anti-retrovirais e aos subtipos genéticos é importante para o controle e tratamento da infecção pelo HIV-1. A pressão exercida pela terapia anti-retroviral seleciona variantes resistentes com mutações nos genes virais da transcriptase reversa (RT) e da protease (PR). Assim, visando contribuir com as autoridades de saúde pública na perspectiva de planejar a disponibilidade de um tratamento terapêutico, nós descrevemos a variabilidade genética e a prevalência de mutações associadas à resistência aos anti-retrovirais em isolados do HIV-1 de indivíduos infectados no Distrito Federal. Dezenove amostras de RNA do HIV-1 provenientes de um laboratório de Saúde Pública do Distrito Federal foram transcritas reversamente e os cDNAs obtidos foram amplificados por nested PCR. Um fragmento de 297 pb correspondente ao gene completo da protease e outro de 647 pb, correspondente à parte do gene da transcriptase reversa (códons 19 a 234), foram obtidos. O seqüenciamento automático e análise de homologia revelaram a presença de 17 subtipos B e 2 subtipos F1 do HIV-1. As seqüências de aminoácidos foram analisadas com relação à presença de mutações associadas à resistência. Um total de 6 mutações na PR, 2 primárias e 4 secundárias e 8 mutações relacionadas à resistência na RT foram encontradas. Nossos dados sugerem uma elevada prevalência do subtipo B do HIV-1 na população estudada do Distrito Federal, assim como a presença de variantes virais geneticamente resistentes em indivíduos que não respondem ao tratamento.

Variants of human papillomavirus types 53, 58 and 66 identified in Central Brazil.

The present study on molecular characterization of human papillomaviruses occurring in Central Brazil, describes two variants each of HPV-53 and HPV-58 and one variant of HPV-66 detected in samples from smears of women showing cervical intraepithelial neoplasia grade II (CIN II). Samples were assayed by PCR using MY09/ MY11 consensus primers, followed by restriction fragment length polymorphism typing. The five isolates showed atypical restriction fragment length profile and MY09/MY11 L1 PCR products were subsequently sequenced. Isolate Bsb-02 and Bsb-08 showed, respectively, 99% similarity to HPV-58 IS404 and 100% to HPV-58 IS417 previously described in the African Continent. Isolates Bsb-61 and Bsb-63 showed 98% similarity to HPV-53, and isolate Bsb-68, 97% similarity to HPV-66. Amino acid substitutions were found in two samples: one in Bsb-02 (T to N) at position 375 and the other in Bsb-61 (S to C) at position 343. Although all the substitutions in Bsb-68 proved to be silent, this sample showed the highest value of pairwise evolutionary distance (2.05%). In countries such as Brazil, where the virus prevalence is high and ethnicity, as well as socio-demographic characteristics, vary according to different regions, HPV variability must be wider and not yet clearly defined.