Targeting HER2/neu with a fully human IgE to harness the allergic reaction against cancer cells

Breast and ovarian cancer are two of the leading causes of cancer deaths among women in the United States. Overexpression of the HER2/neu oncoprotein has been reported in patients affected with breast and ovarian cancers, and is associated with poor prognosis. To develop a novel targeted therapy for HER2/neu expressing tumors, we have constructed a fully human IgE with the variable regions of the scFv C6MH3-B1 specific for HER2/neu. This antibody was expressed in murine myeloma cells and was properly assembled and secreted. The Fc region of this antibody triggers in vitro degranulation of rat basophilic cells expressing human FcεRI (RBL SX-38) in the presence of murine mammary carcinoma cells that express human HER2/neu (D2F2/E2), but not the shed (soluble) antigen (ECD(HER2)) alone. This IgE is also capable of inducing passive cutaneous anaphylaxis in a human FcεRIα transgenic mouse model, in the presence of a cross-linking antibody, but not in the presence of soluble ECD(HER2). Additionally, IgE enhances antigen presentation in human dendritic cells and facilitates cross-priming, suggesting that the antibody is able to stimulate a secondary T-cell anti-tumor response. Furthermore, we show that this IgE significantly prolongs survival of human FcεRIα transgenic mice bearing D2F2/E2 tumors. We also report that the anti-HER2/neu IgE is well tolerated in a preliminary study conducted in Macaca fascicularis (cynomolgus) monkeys. In summary, our results suggest that this IgE should be further explored as a potential therapeutic against HER2/neu overexpressing tumors, such as breast and ovarian cancers.     

Genetic and symbiotic diversity of nitrogen-fixing bacteria isolated from agricultural soils in the Western Amazon by using cowpea as the trap plant

Cowpea is a legume of great agronomic importance that establishes symbiotic relationships with nitrogen-fixing bacteria. However, little is known about the genetic and symbiotic diversity of these bacteria in distinct ecosystems. Our study evaluated the genetic diversity and symbiotic efficiencies of 119 bacterial strains isolated from agriculture soils in the western Amazon using cowpea as a trap plant. These strains were clustered into 11 cultural groups according to growth rate and pH. The 57 nonnodulating strains were predominantly fast growing and acidifying, indicating a high incidence of endophytic strains in the nodules. The other 62 strains, authenticated as nodulating bacteria, exhibited various symbiotic efficiencies, with 68% of strains promoting a significant increase in shoot dry matter of cowpea compared with the control with no inoculation and low levels of mineral nitrogen. Fifty genotypes with 70% similarity and 21 genotypes with 30% similarity were obtained through repetitive DNA sequence (BOX element)-based PCR (BOX-PCR) clustering. The 16S rRNA gene sequencing of strains representative of BOX-PCR clusters showed a predominance of bacteria from the genus Bradyrhizobium but with high species diversity. Rhizobium, Burkholderia, and Achromobacter species were also identified. These results support observations of cowpea promiscuity and demonstrate the high symbiotic and genetic diversity of rhizobia species in areas under cultivation in the western Amazon.     

Encholirium pulchrum and E. diamantinum spp. nov. (Bromeliaceae) from the Espinhaço Range,Minas Gerais,Brazil

Two new species of Encholirium from the Espinhaço range, restricted to the Diamantina plateau of Minas Gerais state, Encholirium pulchrum Forzza, Leme & O. B. C. Ribeiro and Encholirium diamantinum Forzza, are described and illustrated, and their morphological characters are discussed and compared to those of close relatives.     

PyK2 and FAK connections to p190Rho guanine nucleotide exchange factor regulate RhoA activity, focal adhesion formation, and cell motility

Integrin binding to matrix proteins such as fibronectin (FN) leads to formation of focal adhesion (FA) cellular contact sites that regulate migration. RhoA GTPases facilitate FA formation, yet FA-associated RhoA-specific guanine nucleotide exchange factors (GEFs) remain unknown. Here, we show that proline-rich kinase-2 (Pyk2) levels increase upon loss of focal adhesion kinase (FAK) in mouse embryonic fibroblasts (MEFs). Additionally, we demonstrate that Pyk2 facilitates deregulated RhoA activation, elevated FA formation, and enhanced cell proliferation by promoting p190RhoGEF expression. In normal MEFs, p190RhoGEF knockdown inhibits FN-associated RhoA activation, FA formation, and cell migration. Knockdown of p190RhoGEF-related GEFH1 does not affect FA formation in FAK^−/− or normal MEFs. p190RhoGEF overexpression enhances RhoA activation and FA formation in MEFs dependent on FAK binding and associated with p190RhoGEF FA recruitment and tyrosine phosphorylation. These studies elucidate a compensatory function for Pyk2 upon FAK loss and identify the FAK–p190RhoGEF complex as an important integrin-proximal regulator of FA formation during FN-stimulated cell motility.     

Angiosperm epiphytes as conservation indicators in forest fragments: A case study from southeastern Minas Gerais, Brazil

Epiphytes play an important role in the flora and ecology of the tropical regions. Most floristic studies within the state of Minas Gerais focus on terrestrial, woody plant diversity, but this is a different approach, looking at epiphytic angiosperms in three highland areas in Southeastern Minas Gerais, namely Mata do Baú (MB), Reserva Biológica da Represa do Grama (RBRG) and Parque Estadual de Ibitipoca (PEI). Regular collections were performed in these sites between 1999 and 2007, complemented by herbarium records for PEI, and 181 species of epiphytic angiosperms were recorded in 66 genera, within 12 families. Orchidaceae, with 89 species, was by far the largest, and the most species rich genera were Peperomia (Piperaceae) and Pleurothallis s.l. (Orchidaceae), with 12 species each. Similarity analysis has shown a closer relationship between MB and RBRG, both composed by seasonal semideciduous forest, however, Jaccard (0.163) index are low. A similarity analysis including other 21 areas of southeastern and southern Brazil revealed strong influence of sazonality, vegetation type and altitude in the composition of the epiphytic flora, and a relative independence regarding the geographic proximity of the areas sampled. From the conservation standpoint, 30 species recorded for this work appear in the red list for the state of Minas Gerais, under different conservation categories. The low similarity indices obtained between the studied areas underline the importance of the conservation of each one of the remaining forest fragments in Minas Gerais, as their relative geographical proximity does not necessarily mean that their epiphytic flora is similar.     

Highly Pathogenic Avian Influenza Virus Infection of Mallards with Homo- and Heterosubtypic Immunity Induced by Low Pathogenic Avian Influenza Viruses

The potential role of wild birds as carriers of highly pathogenic avian influenza virus (HPAIV) subtype H5N1 is still a matter of debate. Consecutive or simultaneous infections with different subtypes of influenza viruses of low pathogenicity (LPAIV) are very common in wild duck populations. To better understand the epidemiology and pathogenesis of HPAIV H5N1 infections in natural ecosystems, we investigated the influence of prior infection of mallards with homo- (H5N2) and heterosubtypic (H4N6) LPAIV on exposure to HPAIV H5N1. In mallards with homosubtypic immunity induced by LPAIV infection, clinical disease was absent and shedding of HPAIV from respiratory and intestinal tracts was grossly reduced compared to the heterosubtypic and control groups (mean GEC/100 µl at 3 dpi: 3.0×10² vs. 2.3×10⁴ vs. 8.7×10⁴; p<0.05). Heterosubtypic immunity induced by an H4N6 infection mediated a similar but less pronounced effect. We conclude that the epidemiology of HPAIV H5N1 in mallards and probably other aquatic wild bird species is massively influenced by interfering immunity induced by prior homo- and heterosubtypic LPAIV infections.     

Nested in chaos: Insights on the relations of the ‘Nidularioid Complex’ and the evolutionary history of <Emphasis Type="Italic">Neoregelia</Emphasis> (Bromelioideae-Bromeliaceae)

The ‘Nidularioid Complex’ is a group within the Bromelioideae usually characterized by inflorescences nested within water-impounding foliar rosettes. Currently, it comprises six genera: Canistrum, Canistropsis, Edmundoa, Neoregelia, Nidularium, and Wittrockia. While most of these genera occur in eastern Brazil, the distribution of Neoregelia is disjunct between the Atlantic Rainforest and Amazonia. Previous phylogenetic studies have not addressed the monophyly of and relationships among these genera; therefore, we undertook a phylogenetic study of the Nidularioid Complex with emphasis on the genus Neoregelia and its subgenera. A parsimony-based phylogenetic analysis with 101 morphological characters retrieved the Nidularioid Complex as non-monophyletic. Nidularium and Edmundoa were monophyletic. Neoregelia was recovered as non-monophyletic due to the inclusion within it of the Amazonian subgenus Hylaeaicum. These results highlight the need for revision of the generic classification of Bromelioideae, pending increased sampling of taxa and characters.     

Genetic characterization of residual <Emphasis Type="Italic">Triatoma infestans</Emphasis> populations from Brazil by microsatellite

In spite of long-term efforts to eliminate Triatoma infestans (Klug 1834) from Brazil, residual foci still persist in the states of Bahia and Rio Grande do Sul. Data on the genetic variability and structuring of these populations are however lacking. Using nine microsatellite loci, we characterized one residual T. infestans population from Bahia and four from Rio Grande do Sul, and compared them with bugs originally from an older focus in São Paulo; 224 bugs were genotyped. The number of alleles per locus ranged from 5 to 11. Observed and expected heterozygosities per locus ranged, respectively, from 0 to 0.786 and from 0 to 0.764. Significant departures from Hardy–Weinberg equilibrium, mainly due to heterozygote deficits, were detected in all loci and in most populations. Global indices estimated by AMOVA were: Fis was 0.37; Fst was 0.28; and Fit was 0.55; overall indices with p?=?0.00 indicated substantial differentiation. Inter-population Fst ranged from 0.118 to 0.562, suggesting strong genetic structuring and little to no gene flow among populations. Intra-population Fis ranged from 0.301 to 0.307. Inbreeding was apparent in all populations except that from Bahia—which might be either linked by gene flow to nearby unsampled populations or part of a relatively large local population. The overall pattern of strong genetic structuring among pyrethroid-susceptible residual T. infestans populations suggests that their persistence is probably due to operational control failures. Detection and elimination of such residual foci is technically feasible and must become a public health priority in Brazil.