A Bivalent, Spherical Virus-Like Particle Vaccine Enhances Breadth of Immune Responses against Pathogenic Ebola Viruses in Rhesus Macaques.

The 2013-2016 Ebola outbreak in West Africa led to accelerated efforts to develop vaccines against these highly virulent viruses. A live, recombinant vesicular stomatitis virus-based vaccine has been deployed in outbreak settings and appears highly effective. Vaccines based on replication-deficient adenovirus vectors either alone or in combination with a multivalent modified vaccinia Ankara (MVA) Ebola vaccine also appear promising and are progressing in clinical evaluation. However, the ability of current live vector-based approaches to protect against multiple pathogenic species of Ebola is not yet established, and eliciting durable responses may require additional booster vaccinations. Here, we report the development of a bivalent, spherical Ebola virus-like particle (VLP) vaccine that incorporates glycoproteins (GPs) from Zaire Ebola virus (EBOV) and Sudan Ebola virus (SUDV) and is designed to extend the breadth of immunity beyond EBOV. Immunization of rabbits with bivalent Ebola VLPs produced antibodies that neutralized all four pathogenic species of Ebola viruses and elicited antibody-dependent cell-mediated cytotoxicity (ADCC) responses against EBOV and SUDV. Vaccination of rhesus macaques with bivalent VLPs generated strong humoral immune responses, including high titers of binding, as well as neutralizing antibodies and ADCC responses. VLP vaccination led to a significant increase in the frequency of Ebola GP-specific CD4 and CD8 T cell responses. These results demonstrate that a novel bivalent Ebola VLP vaccine elicits strong humoral and cellular immune responses against pathogenic Ebola viruses and support further evaluation of this approach as a potential addition to Ebola vaccine development efforts.IMPORTANCE Ebola outbreaks result in significant morbidity and mortality in affected countries. Although several leading candidate Ebola vaccines have been developed and advanced in clinical testing, additional vaccine candidates may be needed to provide protection against different Ebola species and to extend the durability of protection. A novel approach demonstrated here is to express two genetically diverse glycoproteins on a spherical core, generating a vaccine that can broaden immune responses against known pathogenic Ebola viruses. This approach provides a new method to broaden and potentially extend protective immune responses against Ebola viruses.

Mucosal AIDS virus transmission is enhanced by antiviral IgG isolated early in infection.

OBJECTIVE: Antibody-dependent enhancement (ADE) affects host-virus dynamics in fundamentally different ways: i) enhancement of initial virus acquisition, and/or ii) increased disease progression/severity. Here we address the question whether anti-HIV-1 antibodies can enhance initial infection. While cell-culture experiments hinted at this possibility, in-vivo proof remained elusive. DESIGN: We used passive immunization in nonhuman primates challenged with simian-human immunodeficiency virus (SHIV), a chimera expressing HIV-1 envelope. We purified IgG from rhesus monkeys with early-stage SHIV infection - before cross-neutralizing anti-HIV-1 antibodies had developed - and screened for maximal complement-mediated antibody-dependent enhancement (C'-ADE) of viral replication with a SHIV strain phylogenetically distinct from that harbored by IgG donor macaques. IgG fractions with maximal C'-ADE but lacking neutralization were combined to yield enhancing anti-SHIV IgG (enSHIVIG). RESULTS: We serially enrolled naive macaques (Group 1) to determine the minimal and 50% animal infectious doses required to establish persistent infection after intrarectal SHIV challenge. The first animal was inoculated with a 1 : 10 virus-stock dilution; after this animal's viral RNA load was >104copies/ml, the next macaque was challenged with 10x less virus, a process repeated until viremia no longer ensued. Group 2 was pretreated intravenously with enSHIVIG 24 h before SHIV challenge. Overall, Group 2 macaques required 3.4-fold less virus compared to controls (P = 0.002). This finding is consistent with enhanced susceptibility of the passively immunized animals to mucosal SHIV challenge. CONCLUSION: These passive immunization data give proof of IgG-mediated enhanced virus acquisition after mucosal exposure - a potential concern for antibody-based AIDS vaccine development.

Mucosal Antibodies: Defending Epithelial Barriers against HIV-1 Invasion.

The power of mucosal anti-HIV-1 envelope immunoglobulins (Igs) to block virus transmission is underappreciated. We used passive immunization, a classical tool to unequivocally prove whether antibodies are protective. We mucosally instilled recombinant neutralizing monoclonal antibodies (nmAbs) of different Ig classes in rhesus macaques (RMs) followed by mucosal simian-human immunodeficiency virus (SHIV) challenge. We gave anti-HIV-1 IgM, IgG, and dimeric IgA (dIgA) versions of the same human nmAb, HGN194 that targets the conserved V3 loop crown. Surprisingly, dIgA1 with its wide-open, flat hinge protected 83% of the RMs against intrarectal R5-tropic SHIV-1157ipEL-p challenge, whereas dIgA2, with its narrow hinge, only protected 17% of the animals-despite identical epitope specificities and in vitro neutralization curves of the two dIgA isotypes (Watkins et al., AIDS 2013 27(9):F13-20). These data imply that factors in addition to neutralization determine in vivo protection. We propose that this underlying protective mechanism is immune exclusion, which involves large nmAb/virion aggregates that prevent virus penetration of mucosal barriers. Future studies need to find biomarkers that predict effective immune exclusion in vivo. Vaccine development strategies against HIV-1 and/or other mucosally transmissible pathogens should include induction of strong mucosal Abs of different Ig classes to defend epithelial barriers against pathogen invasion.

A novel Ebola virus antibody-dependent cell-mediated cytotoxicity (Ebola ADCC) assay.

Ebolaviruses are highly virulent pathogens that cause Ebola viral disease (EVD). Data from non-human primate (NHP) models and from human survivors of EVD suggest that anti-Ebola antibodies play an integral role in protection. Antibody-dependent cell-mediated cytotoxicity (ADCC) is a potential mechanism through which anti-Ebola antibodies may mediate protection. We developed a robust Ebola-specific ADCC assay for use in ongoing trials of Ebola vaccines. Stable cell lines for inducible Zaire ebolavirus glycoprotein (EBOV GP) expression were developed to provide a uniform source of target cells in the assay, and were combined with an existing human natural killer (NK) cell line as the effector cell. When applied to commercially available anti-EBOV GP monoclonal antibodies, the assay clearly differentiated antibody with high ADCC activity from those with low or no ADCC activity. Anti-EBOV ADCC activity was also detected in plasma samples from rhesus macaques immunized with a candidate Ebola vaccine. The Ebola ADCC assay reported here will be a useful tool in studying the functionality of anti-EBOV GP antibodies elicited by Ebola vaccines in ongoing and future clinical trials.

Natural Anti-Cancer Agents: Implications in Gemcitabine-Resistant Pancreatic Cancer Treatment.

BACKGROUND & OBJECTIVE: Pancreatic cancer is one of the most lethal malignancies accounting for the fourth leading cause of cancer-related deaths in the United States. Among several explored anticancer agents, Gemcitabine, a nucleoside analogue remained a front line chemotherapeutic agent for the treatment of pancreatic cancer. However, gemcitabine exerts a low response rate with limited progression free survival in patients due to cellular resistance of pancreatic tumors to this therapy. Several chemotherapeutic agents have been explored in combination with gemcitabine against pancreatic cancer with overall mixed responses and survival rates. Naturally occurring dietary agents possess promising anticancer properties and have been shown to target various oncogenic signaling pathways in in-vitro and in-vivo pancreatic cancer models. RESULTS: Multiple studies using natural compounds have shown increased therapeutic efficacy of gemcitabine in pancreatic cancer models. CONCLUSION: This review is focused on recent updates on cellular, preclinical and clinical studies utilizing natural anticancer agents with gemcitabine against pancreatic cancer.