Rapid Induction of Multifunctional Antibodies in Rabbits and Macaques by Clade C HIV-1 CAP257 Envelopes Circulating During Epitope-Specific Neutralization Breadth Development.

We report here on HIV-1 immunization results in rabbits and macaques co-immunized with clade C gp160 DNA and gp140 trimeric envelope vaccines, a strategy similar to a recent clinical trial that showed improved speed and magnitude of humoral responses. Clade C envelopes were isolated from CAP257, an individual who developed a unique temporal pattern of neutralization breadth development, comprising three separate "Waves" targeting distinct Env epitopes and different HIV clades. We used phylogeny and neutralization criteria to down-select envelope vaccine candidates, and confirmed antigenicity of our antigens by interaction with well-characterized broadly neutralizing monoclonal antibodies. Using these envelopes, we performed rabbit studies that screened for immunogenicity of CAP257 Envs from timepoints preceding peak neutralization breadth in each Wave. Selected CAP257 envelopes from Waves 1 and 2, during the first 2 years of infection that were highly immunogenic in rabbits were then tested in macaques. We found that in rabbits and macaques, co-immunization of DNA, and protein envelope-based vaccines induced maximum binding and neutralizing antibody titers with three immunizations. No further benefit was obtained with additional immunizations. The vaccine strategies recapitulated the Wave-specific epitope targeting observed in the CAP257 participant, and elicited Tier 1A, 1B, and Tier 2 heterologous neutralization. CAP257 envelope immunogens also induced the development of ADCC and TFH responses in macaques, and these responses positively correlated with heterologous neutralization. Together, the results from two animal models in this study have implications for identifying effective vaccine immunogens. We used a multi-step strategy to (1) select an Env donor with well-characterized neutralization breadth development; (2) study Env phylogeny for potential immunogens circulating near peak breadth timepoints during the first 2 years of infection; (3) test down-selected Envs for antigenicity; (4) screen down-selected Envs in an effective vaccine regimen in rabbits; and (5) advance the most immunogenic Envs to NHP studies. The results were an induction of high titers of HIV-1 envelope-specific antibodies with increasing avidity and cross-clade neutralizing antibodies with effector functions that together may improve the potential for protection in a pre-clinical SHIV model.

Clinicopathologic Characteristics and Prognosis Analysis of Marjolin's Ulcer in 40 Cases.

Marjolin's ulcer (MU) is a rare but aggressive epidermoid carcinoma observed in scars or wounds. This article provides comprehensive characteristics and prognostic details of MU. Clinical data of 40 patients with MU between January 2010 and December 2017 were analyzed retrospectively. Squamous cell carcinoma was the most common pathological type (35/40, 87.5%). Extended resection was performed to treat all cases with skin grafting or flap grafting. Follow-up duration ranged from 6 to 96 months (median, 52 months) and recurrence was noted in 9 cases. The 1-, 3- and 5-year recurrence-free survival (RFS) rates were 87.2%, 87.2%, 83.2% respectively and the recurrence rate was 22.5%. Univariate analysis revealed that cause of scars (P=0.044), lesion appearance (P=0.036), ultraviolet radiation exposure (P=0.000), depth (P=0.001) and histological grade (P=0.027) had a statistically significant correlation with prognosis of MU. Multivariate analysis revealed that depth (P=0.034, RR=2.681, 95%CI: 1.077-6.674) and histological grade (P=0.008, RR=2.820, 95%CI: 1.315-6.050) were independent prognostic factors for RFS. In conclusion, superficial infiltration and high-grade differentiation predict more favorable prognosis. Careful follow-up of high-risk groups is strongly recommended to prevent recurrence and improve prognosis.

Envelope variants circulating as initial neutralization breadth developed in two HIV-infected subjects stimulate multiclade neutralizing antibodies in rabbits.

UNLABELLED: Identifying characteristics of the human immunodeficiency virus type 1 (HIV-1) envelope that are effective in generating broad, protective antibodies remains a hurdle to HIV vaccine design. Emerging evidence of the development of broad and potent neutralizing antibodies in HIV-infected subjects suggests that founder and subsequent progeny viruses may express unique antigenic motifs that contribute to this developmental pathway. We hypothesize that over the course of natural infection, B cells are programmed to develop broad antibodies by exposure to select populations of emerging envelope quasispecies variants. To test this hypothesis, we identified two unrelated subjects whose antibodies demonstrated increasing neutralization breadth against a panel of HIV-1 isolates over time. Full-length functional env genes were cloned longitudinally from these subjects from months after infection through 2.6 to 5.8 years of infection. Motifs associated with the development of breadth in published, cross-sectional studies were found in both subjects. We compared the immunogenicity of envelope vaccines derived from time points obtained during and after broadening of neutralization activity within these subjects. Rabbits were coimmunized four times with selected multiple gp160 DNAs and gp140-trimeric envelope proteins. The affinity of the polyclonal response increased as a function of boosting. The most rapid and persistent neutralization of multiclade tier 1 viruses was elicited by envelopes that were circulating in plasma at time points prior to the development of 50% neutralization breadth in both human subjects. The breadth elicited in rabbits was not improved by exposure to later envelope variants. These data have implications for vaccine development in describing a target time point to identify optimal envelope immunogens. IMPORTANCE: Vaccine protection against viral infections correlates with the presence of neutralizing antibodies; thus, vaccine components capable of generating potent neutralization are likely to be critical constituents in an effective HIV vaccine. However, vaccines tested thus far have elicited only weak antibody responses and very modest, waning protection. We hypothesized that B cells develop broad antibodies by exposure to the evolving viral envelope population and tested this concept using multiple envelopes from two subjects who developed neutralization breadth within a few years of infection. We compared different combinations of envelopes from each subject to identify the most effective immunogens and regimens. In each subject, use of HIV envelopes circulating during the early development and maturation of breadth generated more-potent antibodies that were modestly cross neutralizing. These data suggest a new approach to identifying envelope immunogens that may be more effective in generating protective antibodies in humans.

Neutralizing polyclonal IgG present during acute infection prevents rapid disease onset in simian-human immunodeficiency virus SHIVSF162P3-infected infant rhesus macaques.

Simian-human immunodeficiency virus (SHIV) models for human immunodeficiency virus (HIV) infection have been widely used in passive studies with HIV neutralizing antibodies (NAbs) to test for protection against infection. However, because SHIV-infected adult macaques often rapidly control plasma viremia and any resulting pathogenesis is minor, the model has been unsuitable for studying the impact of antibodies on pathogenesis in infected animals. We found that SHIVSF162P3 infection in 1-month-old rhesus macaques not only results in high persistent plasma viremia but also leads to very rapid disease progression within 12 to 16 weeks. In this model, passive transfer of high doses of neutralizing IgG (SHIVIG) prevents infection. Here, we show that at lower doses, SHIVIG reduces both plasma and peripheral blood mononuclear cell (PBMC)-associated viremia and mitigates pathogenesis in infected animals. Moreover, production of endogenous NAbs correlated with lower set-point viremia and 100% survival of infected animals. New SHIV models are needed to investigate whether passively transferred antibodies or antibodies elicited by vaccination that fall short of providing sterilizing immunity impact disease progression or influence immune responses. The 1-month-old rhesus macaque SHIV model of infection provides a new tool to investigate the effects of antibodies on viral replication and clearance, mechanisms of B cell maintenance, and the induction of adaptive immunity in disease progression.

Conditions that influence the response to Fgf during otic placode induction.

Despite the vital importance of Fgf for otic induction, previous attempts to study otic induction through Fgf misexpression have yielded widely varying and contradictory results. There are also discrepancies regarding the ability of Fgf to induce otic tissue in ectopic locations, raising questions about the sufficiency of Fgf and the degree to which other local factors enhance or restrict otic potential. Using heat shock-inducible transgenes to misexpress Fgf3 or Fgf8 in zebrafish, we found that the stage, distribution and level of misexpression strongly influence the response to Fgf. Fgf misexpression during gastrulation can inhibit or promote otic development, depending on context, whereas misexpression after gastrulation leads to expansion of otic markers throughout preplacodal ectoderm surrounding the head. Elevated Fgf also expands expression of the putative competence factor Foxi1, which is required for Fgf to expand other otic markers. Misexpression of downstream factors Pax2a or Pax8 also expands otic markers but cannot bypass the requirement for Fgf or Foxi1. Co-misexpression of Pax2/8 with Fgf8 potentiates formation of ectopic otic vesicles expressing a full range of otic markers. These findings document the variables critically affecting the response to Fgf and clarify the roles of foxi1 and pax2/8 in the otic response.

Differential expression and phylogenetic analysis suggest specialization of plastid-localized members of the PHT4 phosphate transporter family for photosynthetic and heterotrophic tissues.

Plastids rely on multiple phosphate (Pi) transport activities to support and control a wide range of metabolic processes, including photosynthesis and carbon partitioning. Five of the six members of the PHT4 family of Pi transporters in Arabidopsis thaliana (PHT4;1-PHT4;5) are confirmed or predicted plastid proteins. As a step towards identifying the roles of individual PHT4 Pi transporters in chloroplast and non-photosynthetic plastid Pi dynamics, we used promoter-reporter gene fusions and quantitative RT-PCR studies, respectively, to determine spatial and diurnal gene expression patterns. PHT4;1 and PHT4;4 were both expressed predominantly in photosynthetic tissues, although expression of PHT4;1 was circadian and PHT4;4 was induced by light. PHT4;3 and PHT4;5 were expressed mainly in leaf phloem. PHT4;2 was expressed throughout the root, and exhibited a diurnal pattern with peak transcript levels in the dark. The remaining member of this gene family, PHT4;6, encodes a Golgi-localized protein and was expressed ubiquitously. The overlapping but distinct expression patterns for these genes suggest specialized roles for the encoded transporters in multiple types of differentiated plastids. Phylogenetic analysis revealed conservation of each of the orthologous members of the PHT4 family in Arabidopsis and rice, which is consistent with specialization, and suggests that the individual members of this transporter family diverged prior to the divergence of monocots and dicots.