Un examen actualizado de la percepción de las barreras para la implementación de la farmacogenómica y la utilidad de los pares fármaco/gen en América Latina y el Caribe

La farmacogenómica (PGx) se considera un campo emergente en los países en desarrollo. La investigación sobre PGx en la región de América Latina y el Caribe (ALC) sigue siendo escasa, con información limitada en algunas poblaciones. Por lo tanto, las extrapolaciones son complicadas, especialmente en poblaciones mixtas. En este trabajo, revisamos y analizamos el conocimiento farmacogenómico entre la comunidad científica y clínica de ALC y examinamos las barreras para la aplicación clínica. Realizamos una búsqueda de publicaciones y ensayos clínicos en este campo en todo el mundo y evaluamos la contribución de ALC. A continuación, realizamos una encuesta regional estructurada que evaluó una lista de 14 barreras potenciales para la aplicación clínica de biomarcadores en función de su importancia. Además, se analizó una lista emparejada de 54 genes/fármacos para determinar una asociación entre los biomarcadores y la respuesta a la medicina genómica. Esta encuesta se comparó con una encuesta anterior realizada en 2014 para evaluar el progreso en la región. Los resultados de la búsqueda indicaron que los países de América Latina y el Caribe han contribuido con el 3,44% del total de publicaciones y el 2,45% de los ensayos clínicos relacionados con PGx en todo el mundo hasta el momento. Un total de 106 profesionales de 17 países respondieron a la encuesta. Se identificaron seis grandes grupos de obstáculos. A pesar de los continuos esfuerzos de la región en la última década, la principal barrera para la implementación de PGx en ALC sigue siendo la misma, la "necesidad de directrices, procesos y protocolos para la aplicación clínica de la farmacogenética/farmacogenómica". Las cuestiones de coste-eficacia se consideran factores críticos en la región. Los puntos relacionados con la reticencia de los clínicos son actualmente menos relevantes. Según los resultados de la encuesta, los pares gen/fármaco mejor clasificados (96%-99%) y percibidos como importantes fueron CYP2D6/tamoxifeno, CYP3A5/tacrolimus, CYP2D6/opioides, DPYD/fluoropirimidinas, TMPT/tiopurinas, CYP2D6/antidepresivos tricíclicos, CYP2C19/antidepresivos tricíclicos, NUDT15/tiopurinas, CYP2B6/efavirenz y CYP2C19/clopidogrel. En conclusión, aunque la contribución global de los países de ALC sigue siendo baja en el campo del PGx, se ha observado una mejora relevante en la región. La percepción de la utilidad de las pruebas PGx en la comunidad biomédica ha cambiado drásticamente, aumentando la concienciación entre los médicos, lo que sugiere un futuro prometedor en las aplicaciones clínicas de PGx en ALC.Pharmacogenomics (PGx) is considered an emergent field in developing countries. Research on PGx in the Latin American and the Caribbean (LAC) region remains scarce, with limited information in some populations. Thus, extrapolations are complicated, especially in mixed populations. In this paper, we reviewed and analyzed pharmacogenomic knowledge among the LAC scientific and clinical community and examined barriers to clinical application. We performed a search for publications and clinical trials in the field worldwide and evaluated the contribution of LAC. Next, we conducted a regional structured survey that evaluated a list of 14 potential barriers to the clinical implementation of biomarkers based on their importance. In addition, a paired list of 54 genes/drugs was analyzed to determine an association between biomarkers and response to genomic medicine. This survey was compared to a previous survey performed in 2014 to assess progress in the region. The search results indicated that Latin American and Caribbean countries have contributed 3.44% of the total publications and 2.45% of the PGx-related clinical trials worldwide thus far. A total of 106 professionals from 17 countries answered the survey. Six major groups of barriers were identified. Despite the region’s continuous efforts in the last decade, the primary barrier to PGx implementation in LAC remains the same, the “need for guidelines, processes, and protocols for the clinical application of pharmacogenetics/pharmacogenomics”. Cost-effectiveness issues are considered critical factors in the region. Items related to the reluctance of clinicians are currently less relevant. Based on the survey results, the highest ranked (96%–99%) gene/drug pairs perceived as important were CYP2D6/tamoxifen, CYP3A5/tacrolimus, CYP2D6/opioids, DPYD/fluoropyrimidines, TMPT/thiopurines, CYP2D6/tricyclic antidepressants, CYP2C19/tricyclic antidepressants, NUDT15/thiopurines, CYP2B6/efavirenz, and CYP2C19/clopidogrel. In conclusion, although the global contribution of LAC countries remains low in the PGx field, a relevant improvement has been observed in the region. The perception of the usefulness of PGx tests in biomedical community has drastically changed, raising awareness among physicians, which suggests a promising future in the clinical applications of PGx in LAC

Age-related immune response heterogeneity to SARS-CoV-2 vaccine BNT162b2

Abstract: Although two-dose mRNA vaccination provides excellent protection against SARS-CoV-2, there is little information about vaccine efficacy against variants of concern (VOC) in individuals above eighty years of age1. Here we analysed immune responses following vaccination with the BNT162b2 mRNA vaccine2 in elderly participants and younger healthcare workers. Serum neutralization and levels of binding IgG or IgA after the first vaccine dose were lower in older individuals, with a marked drop in participants over eighty years old. Sera from participants above eighty showed lower neutralization potency against the B.1.1.7 (Alpha), B.1.351 (Beta) and P.1. (Gamma) VOC than against the wild-type virus and were more likely to lack any neutralization against VOC following the first dose. However, following the second dose, neutralization against VOC was detectable regardless of age. The frequency of SARS-CoV-2 spike-specific memory B cells was higher in elderly responders (whose serum showed neutralization activity) than in non-responders after the first dose. Elderly participants showed a clear reduction in somatic hypermutation of class-switched cells. The production of interferon-γ and interleukin-2 by SARS-CoV-2 spike-specific T cells was lower in older participants, and both cytokines were secreted primarily by CD4 T cells. We conclude that the elderly are a high-risk population and that specific measures to boost vaccine responses in this population are warranted, particularly where variants of concern are circulating

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

High polymorphism in genes encoding antigen B from human infecting strains of Echinococcus granulosus

Echinococcus granulosus antigen B (AgB) is encoded by a gene family and is involved in the evasion of the host immune response. E. granulosus exists as a number of strains (G1–G10) that differ in biological characteristics. We used PCR-SSCP followed by DNA sequencing to evaluate sequence variation and transcription profile of AgB in 5 E. granulosus strains#. Twenty-four genomic sequences were isolated and clustered in 3 groups related to 2 of the 5 reported AgB genes. AgB4 genes were present in almost all strains, whereas AgB2 were present as functional genes exclusively in G1/G2 cluster, and as non-functional genes in G5 and the G6/G7 cluster, suggesting inter-strain variation The AgB transcription patterns, analysed by RT-PCR, showed that AgB2 and AgB4 genes were transcribed in G1, while only the AgB4 gene was transcribed in G7 strain. Cysts from the same strain or cluster shared more genomic and cDNA variants than cysts from different strain or cluster. The level of nucleotide and deduced amino acid sequence variation observed is higher than that reported so far for coding genes of other helminths. Neutrality was rejected for AgB2 genes. These data show the genetic polymorphism of antigen-coding genes among genetically characterized strains of E. granulosus

High polymorphism in genes encoding antigen B from human infecting strains of Echinococcus granulosus

Echinococcus granulosus antigen B (AgB) is encoded by a gene family and is involved in the evasion of the host immune response. E. granulosus exists as a number of strains (G1–G10) that differ in biological characteristics. We used PCR-SSCP followed by DNA sequencing to evaluate sequence variation and transcription profile of AgB in 5 E. granulosus strains#. Twenty-four genomic sequences were isolated and clustered in 3 groups related to 2 of the 5 reported AgB genes. AgB4 genes were present in almost all strains, whereas AgB2 were present as functional genes exclusively in G1/G2 cluster, and as non-functional genes in G5 and the G6/G7 cluster, suggesting inter-strain variation The AgB transcription patterns, analysed by RT-PCR, showed that AgB2 and AgB4 genes were transcribed in G1, while only the AgB4 gene was transcribed in G7 strain. Cysts from the same strain or cluster shared more genomic and cDNA variants than cysts from different strain or cluster. The level of nucleotide and deduced amino acid sequence variation observed is higher than that reported so far for coding genes of other helminths. Neutrality was rejected for AgB2 genes. These data show the genetic polymorphism of antigen-coding genes among genetically characterized strains of E. granulosus

Sequencing, bioinformatic characterization and expression pattern of a putative amino acid transporter from the parasitic cestode Echinococcus granulosus

Fil: Camicia, Federico. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Paredes, Rodolfo. Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Independencia 1027, Santiago; ChileFil: Chalar, Cora. Sección Bioquímica, Instituto de Biología, Facultad de Ciencias, Universidad de la República, Iguá 4225, Montevideo; Uruguay.Fil: Galanti, Norbel. Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Independencia 1027, Santiago; ChileFil: Kamenetzky, Laura. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Gutierrez, Ariana. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Rosenzvit, Mara C. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.We have sequenced and partially characterized an Echinococcus granulosus cDNA, termed egat1, from a protoscolex signal sequence trap (SST) cDNA library. The isolated 1627 bp long cDNA contains an ORF of 489 amino acids and shows an amino acid identity of 30% with neutral and excitatory amino acid transporters members of the Dicarboxylate/Amino Acid Na+ and/or H+ Cation Symporter family (DAACS) (TC 2.A.23). Additional bioinformatics analysis of EgAT1, confirmed the results obtained by similarity searches and showed the presence of 9 to 10 transmembrane domains, consensus sequences for N-glycosylation between the third and fourth transmembrane domain, a highly similar hydropathy profile with ASCT1 (a known member of DAACS family), high score with SDF (Sodium Dicarboxilate Family) and similar motifs with EDTRANSPORT, a fingerprint of excitatory amino acid transporters. The localization of the putative amino acid transporter was analyzed by in situ hybridization and immunofluorescence in protoscoleces and associated germinal layer. The in situ hybridization labelling indicates the distribution of egat1 mRNA throughout the tegument. EgAT1 protein, which showed in Western blots a molecular mass of approximately 60 kD, is localized in the subtegumental region of the metacestode, particularly around suckers and rostellum of protoscoleces and layers from brood capsules. The sequence and expression analyses of EgAT1 pave the way for functional analysis of amino acids transporters of E. granulosus and its evaluation as new drug targets against cystic echinococcosis

High polymorphism in genes encoding antigen B from human infecting strains of Echinococcus granulosus

Echinococcus granulosus antigen B (AgB) is encoded by a gene family and is involved in the evasion of the host immune response. E. granulosus exists as a number of strains (G1–G10) that differ in biological characteristics. We used PCR-SSCP followed by DNA sequencing to evaluate sequence variation and transcription profile of AgB in 5 E. granulosus strains#. Twenty-four genomic sequences were isolated and clustered in 3 groups related to 2 of the 5 reported AgB genes. AgB4 genes were present in almost all strains, whereas AgB2 were present as functional genes exclusively in G1/G2 cluster, and as non-functional genes in G5 and the G6/G7 cluster, suggesting inter-strain variation The AgB transcription patterns, analysed by RT-PCR, showed that AgB2 and AgB4 genes were transcribed in G1, while only the AgB4 gene was transcribed in G7 strain. Cysts from the same strain or cluster shared more genomic and cDNA variants than cysts from different strain or cluster. The level of nucleotide and deduced amino acid sequence variation observed is higher than that reported so far for coding genes of other helminths. Neutrality was rejected for AgB2 genes. These data show the genetic polymorphism of antigen-coding genes among genetically characterized strains of E. granulosus

High polymorphism in genes encoding antigen B from human infecting strains of Echinococcus granulosus

Fil: Kamenetzky, Laura. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Muzulin, Patricia Marcela. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Gutiérrez, Ariana M. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Angel, Sergio O. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto Tecnológico de Chascomús; Argentina.Fil: Zaha, A. Universidade Federal do Rio Grande do Sul. Instituto de Biociências. Centro de Biotecnologia e Departamento de Biologia Molecular e Biotecnologia; Brasil.Fil: Guarnera, Eduardo A. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Rosenzvit, Mara C. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Echinococcus granulosus antigen B (AgB) is encoded by a gene family and is involved in the evasion of the host immune response. E. granulosus exists as a number of strains (G1-G10) that differ in biological characteristics. We used PCR-SSCP followed by DNA sequencing to evaluate sequence variation and transcription profile of AgB in 5 E. granulosus strains. Twenty-four genomic sequences were isolated and clustered in 3 groups related to 2 of the 5 reported AgB genes. AgB4 genes were present in almost all strains, whereas AgB2 were present as functional genes exclusively in G1/G2 cluster, and as non-functional genes in G5 and the G6/G7 cluster, suggesting inter-strain variation. The AgB transcription patterns, analysed by RT-PCR, showed that AgB2 and AgB4 genes were transcribed in G1, while only the AgB4 gene was transcribed in G7 strain. Cysts from the same strain or cluster shared more genomic and cDNA variants than cysts from different strain or cluster. The level of nucleotide and deduced amino acid sequence variation observed is higher than that reported so far for coding genes of other helminths. Neutrality was rejected for AgB2 genes. These data show the genetic polymorphism of antigen-coding genes among genetically characterized strains of E. granulosus

Patent and pre-patent detection of Echinococcus granulosus genotypes in the definitive host

Fil: Naidich, Ariel. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: McManus, Donald P. Royal Brisbane Hospital Post Office. Queensland Institute of Medical Research. Molecular Parasitology Laboratory. Division of Infectious Diseases and Immunology; Australia.Fil: Canova, Sergio G. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Gutierrez, Ariana M. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Zhang, Wenbao. Royal Brisbane Hospital Post Office. Queensland Institute of Medical Research. Molecular Parasitology Laboratory. Division of Infectious Diseases and Immunology; Australia.Fil: Guarnera, Eduardo A. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Rosenzvit, Mara Cecilia. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.The detection of Echinococcus granulosus in dogs is important for epidemiological surveillance and evaluation of cystic hydatic disease control programs. We report the efficacy of two PCR-based methods to detect patent and pre-patent infection in dogs experimentally infected with E. granulosus. The detection is based on amplification of a fragment of a mitochondrial gene (Mit-PCR) and a DNA repetitive element (Rep-PCR) of E. granulosus. We tested the ability of both methods to detect several genotypes of the parasite. Both PCR methods could detect E. granulosus in pre-patent and patent periods, even when microscopical observation of eggs resulted negative in fecal samples. The Mit-PCR produced the same amplification pattern for all the parasite genotypes tested while the amplification patterns with the Rep-PCR differed among groups of strains. Fecal samples collected from dogs of an endemic area were diagnosed with more sensitivity than arecoline hydrobromide purgation. These molecular methods could be applied in the confirmation of coproantigen-positive fecal samples and to verify the success of control programs

High polymorphism in genes encoding antigen B from human infecting strains of Echinococcus granulosus

Fil: Kamenetzky, Laura. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Muzulin, Patricia Marcela. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Gutiérrez, Ariana M. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Angel, Sergio O. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto Tecnológico de Chascomús; Argentina.Fil: Zaha, A. Universidade Federal do Rio Grande do Sul. Instituto de Biociências. Centro de Biotecnologia e Departamento de Biologia Molecular e Biotecnologia; Brasil.Fil: Guarnera, Eduardo A. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Rosenzvit, Mara C. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Echinococcus granulosus antigen B (AgB) is encoded by a gene family and is involved in the evasion of the host immune response. E. granulosus exists as a number of strains (G1-G10) that differ in biological characteristics. We used PCR-SSCP followed by DNA sequencing to evaluate sequence variation and transcription profile of AgB in 5 E. granulosus strains. Twenty-four genomic sequences were isolated and clustered in 3 groups related to 2 of the 5 reported AgB genes. AgB4 genes were present in almost all strains, whereas AgB2 were present as functional genes exclusively in G1/G2 cluster, and as non-functional genes in G5 and the G6/G7 cluster, suggesting inter-strain variation. The AgB transcription patterns, analysed by RT-PCR, showed that AgB2 and AgB4 genes were transcribed in G1, while only the AgB4 gene was transcribed in G7 strain. Cysts from the same strain or cluster shared more genomic and cDNA variants than cysts from different strain or cluster. The level of nucleotide and deduced amino acid sequence variation observed is higher than that reported so far for coding genes of other helminths. Neutrality was rejected for AgB2 genes. These data show the genetic polymorphism of antigen-coding genes among genetically characterized strains of E. granulosus