Efficacy and breadth of adjuvanted SARS-CoV-2 receptor-binding domain nanoparticle vaccine in macaques

Emergence of novel variants of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) underscores the need for next-generation vaccines able to elicit broad and durable immunity. Here we report the evaluation of a ferritin nanoparticle vaccine displaying the receptor-binding domain of the SARS-CoV-2 spike protein (RFN) adjuvanted with Army Liposomal Formulation QS-21 (ALFQ). RFN vaccination of macaques using a two-dose regimen resulted in robust, predominantly Th1 CD4+ T cell responses and reciprocal peak mean serum neutralizing antibody titers of 14,000 to 21,000. Rapid control of viral replication was achieved in the upper and lower airways of animals after high-dose SARS-CoV-2 respiratory challenge, with undetectable replication within 4 d in seven of eight animals receiving 50 µg of RFN. Cross-neutralization activity against SARS-CoV-2 variant B.1.351 decreased only approximately twofold relative to WA1/2020. In addition, neutralizing, effector antibody and cellular responses targeted the heterotypic SARS-CoV-1, highlighting the broad immunogenicity of RFN-ALFQ for SARS-CoV−like Sarbecovirus vaccine development.

Induction of Protection in Mice against a Chlamydia muridarum Respiratory Challenge by a Vaccine Formulated with the Major Outer Membrane Protein in Nanolipoprotein Particles

Chlamydia trachomatis is a sexually transmitted bacterium that infects over 130 million individuals worldwide annually. To implement a vaccine, we developed a cell-free co-translational system to express the Chlamydia muridarum major outer membrane protein (MOMP). This approach uses a nanolipoprotein particles (tNLP) made from ApoA1 protein, amphiphilic telodendrimer and lipids that self-assemble to form 10–25 nm discs. These tNLP provide a protein-encapsulated lipid support to solubilize and fold membrane proteins. The cell-free system co-translated MOMP and ApoA1 in the presence of telodendrimer mixed with lipids. The MOMP-tNLP complex was amenable to CpG and FSL-1 adjuvant addition. To investigate the ability of MOMP-tNLP+CpG+FSL-1 to induce protection against an intranasal (i.n.) C. muridarum challenge, female mice were vaccinated intramuscularly (i.m.) or i.n. and i.m. simultaneously 4 weeks apart. Following vaccination with MOMP-tNLP+CpG+FSL-1, mice mounted significant humoral and cell-mediated immune responses. Following the i.n. challenge, mice vaccinated with MOMP-tNLP+CpG+FSL-1 i.n. + i.m. group were protected as determined by the percentage change in body weight and by the number of C. muridarum inclusion forming units (IFU) recovered from the lungs. To our knowledge, this is the first time a MOMP-based vaccine formulated in tNLP has been shown to protect against C. muridarum.

Respiratory dysfunction in a mouse model of spinocerebellar ataxia type 7

Spinocerebellar ataxia type 7 (SCA7) is an autosomal dominant neurodegenerative disorder caused by a CAG repeat expansion in the coding region of the ataxin-7 gene. Infantile-onset SCA7 patients display extremely large repeat expansions (>200 CAGs) and exhibit progressive ataxia, dysarthria, dysphagia and retinal degeneration. Severe hypotonia, aspiration pneumonia and respiratory failure often contribute to death in affected infants. To better understand the features of respiratory and upper airway dysfunction in SCA7, we examined breathing and putative phrenic and hypoglossal neuropathology in a knock-in mouse model of early-onset SCA7 carrying an expanded allele with 266 CAG repeats. Whole-body plethysmography was used to measure awake, spontaneous breathing at baseline in normoxia and during a hypercapnic/hypoxic respiratory challenge at 4 and 8 weeks, before and after onset of disease. Postmortem studies included quantification of putative phrenic and hypoglossal motor neurons and microglia and analysis of ataxin-7 aggregation at end stage. SCA7-266Q mice have profound breathing deficits during a respiratory challenge, exhibiting reduced respiratory output and a greater percentage of time in apnea. Histologically, putative phrenic and hypoglossal motor neurons of SCA7 mice exhibit a reduction in number accompanied by increased microglial activation, indicating neurodegeneration and neuroinflammation. Furthermore, intranuclear ataxin-7 accumulation is observed in cells neighboring putative phrenic and hypoglossal motor neurons in SCA7 mice. These findings reveal the importance of phrenic and hypoglossal motor neuron pathology associated with respiratory failure and upper airway dysfunction, which are observed in infantile-onset SCA7 patients and likely contribute to their early death.

Immunogenicity and protectivity of the peptide candidate vaccine against SARS-CoV-2

Background: In 2020, the pandemic caused by novel coronavirus infection has become one of the most critical global health challenges during the past century. The lack of a vaccine, as the most effective way to control the novel infection, has prompted the development of a large number of preventive products by the scientific community. We have developed a candidate vaccine (EpiVacCorona) against novel coronavirus infection caused by SARS-CoV-2 that is based on chemically synthesized peptides conjugated to a carrier protein and adsorbed on aluminum hydroxide and studied the specific activity of the developed vaccine. Aims: Study of the immunogenicity and protectivity of the peptide candidate vaccine EpiVacCorona. Materials and methods: the work was performed using standard molecular biological, virological and histological methods. Results: It was demonstrated that EpiVacCorona, when administered twice, spaced 14 days apart, to hamsters, ferrets, and non-human primates (African green monkeys, rhesus macaques) at a dose of 260 g, which is equal to one inoculation dose for humans, induces virus-specific antibodies in 100% of the animals. Experiments in hamsters showed this vaccine to be associated with the dose-dependent immunogenicity. The vaccine was shown to accelerate the elimination of the virus from the upper respiratory tract in ferrets and prevent the development of pneumonia in hamsters and non-human primates following a respiratory challenge with novel coronavirus. Conclusions: The results of a preclinical specific activity study indicate that the use of EpiVacCorona has the potential for human vaccination.

Efficacy of a Broadly Neutralizing SARS-CoV-2 Ferritin Nanoparticle Vaccine in Nonhuman Primates

The emergence of novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants stresses the continued need for next-generation vaccines that confer broad protection against coronavirus disease 2019 (COVID-19). We developed and evaluated an adjuvanted SARS-CoV-2 Spike Ferritin Nanoparticle (SpFN) vaccine in nonhuman primates (NHPs). High-dose (50µg) SpFN vaccine, given twice within a 28 day interval, induced a Th1-biased CD4 T cell helper response and a peak neutralizing antibody geometric mean titer of 52,773 against wild-type virus, with activity against SARS-CoV-1 and minimal decrement against variants of concern. Vaccinated animals mounted an anamnestic response upon high-dose SARS-CoV-2 respiratory challenge that translated into rapid elimination of replicating virus in their upper and lower airways and lung parenchyma. SpFN’s potent and broad immunogenicity profile and resulting efficacy in NHPs supports its utility as a candidate platform for SARS-like betacoronaviruses.One-Sentence SummaryA SARS-CoV-2 Spike protein ferritin nanoparticle vaccine, co-formulated with a liposomal adjuvant, elicits broad neutralizing antibody responses that exceed those observed for other major vaccines and rapidly protects against respiratory infection and disease in the upper and lower airways and lung tissue of nonhuman primates.

Zinc Source and Concentration Altered Physiological Responses of Beef Heifers during a Combined Viral-Bacterial Respiratory Challenge

To determine the effects of zinc supplementation on the immune response to a combined viral-bacterial respiratory disease challenge, thirty-two beef heifers (255 ± 15 kg) were subjected to a 30-d period of Zn depletion, then randomly assigned to one of three treatment diets fed for 30 d before the challenge: (1) supplementation with 100 mg of Zn from Zn sulfate/kg of DM (Zn100), (2) supplementation with 200 mg of Zn from Zn sulfate/kg of DM (Zn200), and (3) supplementation with 80 mg of Zn/kg of DM from zinc methionine and 20 mg of Zn from Zn sulfate/kg of DM (ZinMet). After the 30-d supplementation period, all heifers were fitted with indwelling vaginal temperature (VT) devices and intra-nasally challenged with 1 × 108 PFU bovine herpesvirus-1 on d -3, and then allowed to rest in outdoor pens for 3 d. On d 0, each heifer was challenged intra-tracheally with an average dose of 2.38 × 107 CFU Mannheimia haemolytica (MH), fitted with an indwelling jugular catheter, and then moved into individual stalls in an environmentally-controlled enclosed barn. Whole blood samples were collected at 1-h (serum) and 2-h (complete blood counts) intervals from 0 to 8 h, and at 12, 24, 36, 48, 60, 72, 168, and 360 h relative to MH challenge. Data were analyzed using the MIXED procedure of SAS specific for repeated measures with fixed effects of treatment, time, and their interaction. There was a treatment effect (p < 0.01) for VT such that Zn200 heifers had greater VT than Zn100 and ZinMet heifers. There was a trend (p = 0.10) for a serum cortisol treatment effect with Zn100 heifers having greater cortisol than ZinMet heifers. Total leukocytes and lymphocytes were greater (p ≤ 0.01) in Zn100 heifers than Zn200 and ZinMet heifers, whereas monocytes were less (p = 0.05) in ZinMet heifers than Zn100 and Zn200 heifers. Concentrations of IL-6 were greater (p = 0.02) in ZinMet heifers than Zn100 and Zn200 heifers. Concentrations of IFN-γ were greater in Zn200 heifers than ZinMet heifers at 0 h, and Zn100 heifers from 0 to 12 h post-MH challenge (treatment x time p = 0.02). Serum haptoglobin was not affected by treatment or treatment x time (p ≥ 0.36) but increased over time (p < 0.01) in all groups. There was a trend (p = 0.11) for ZinMet heifers to have less severe nasal lesion scores than Zn100 heifers. The observed differential physiological responses in this study indicate that zinc source and concentration may alter the response to a bovine respiratory challenge in heifers.

Epicutaneous immunization with modified vaccinia Ankara viral vectors generates superior T cell immunity against a respiratory viral challenge

Modified Vaccinia Ankara (MVA) was recently approved as a smallpox vaccine. Variola is transmitted by respiratory droplets and MVA immunization by skin scarification (s.s.) protected mice far more effectively against lethal respiratory challenge with vaccinia virus (VACV) than any other route of delivery, and at lower doses. Comparisons of s.s. with intradermal, subcutaneous, or intramuscular routes showed that MVAOVA s.s.-generated T cells were both more abundant and transcriptionally unique. MVAOVA s.s. produced greater numbers of lung Ova-specific CD8+ TRM and was superior in protecting mice against lethal VACVOVA respiratory challenge. Nearly as many lung TRM were generated with MVAOVA s.s. immunization compared to intra-tracheal immunization with MVAOVA and both routes vaccination protected mice against lethal pulmonary challenge with VACVOVA. Strikingly, MVAOVA s.s.-generated effector T cells exhibited overlapping gene transcriptional profiles to those generated via intra-tracheal immunization. Overall, our data suggest that heterologous MVA vectors immunized via s.s. are uniquely well-suited as vaccine vectors for respiratory pathogens, which may be relevant to COVID-19. In addition, MVA delivered via s.s. could represent a more effective dose-sparing smallpox vaccine.

Cerebral oxygen extraction fraction (OEF): Comparison of challenge-free gradient echo QSM+qBOLD (QQ) with 15O PET in healthy adults

We aimed to validate oxygen extraction fraction (OEF) estimations by quantitative susceptibility mapping plus quantitative blood oxygen-level dependence (QSM+qBOLD, or QQ) using 15O-PET. In ten healthy adult brains, PET and MRI were acquired simultaneously on a PET/MR scanner. PET was acquired using C[15O], O[15O], and H2[15O]. Image-derived arterial input functions and standard models of oxygen metabolism provided quantification of PET. MRI included T1-weighted imaging, time-of-flight angiography, and multi-echo gradient-echo imaging that was processed for QQ. Region of interest (ROI) analyses compared PET OEF and QQ OEF. In ROI analyses, the averaged OEF differences between PET and QQ were generally small and statistically insignificant. For whole brains, the average and standard deviation of OEF was 32.8 ± 6.7% for PET; OEF was 34.2 ± 2.6% for QQ. Bland-Altman plots quantified agreement between PET OEF and QQ OEF. The interval between the 95% limits of agreement was 16.9 ± 4.0% for whole brains. Our validation study suggests that respiratory challenge-free QQ-OEF mapping may be useful for non-invasive clinical assessment of regional OEF impairment.

64 Immunological, physiological, and behavioral responses to an experimentally-induced respiratory disease challenge in growing steers

Immunologic, physiologic, and behavioral responses to a combined viral-bacterial respiratory challenge were explored in beef steers (initial BW 293 kg). Steers (n = 24) were inoculated intranasally with bovine herpes virus-1 (2×108 PFU) and intratracheally with Mannheimia haemolytica (MH, 2.15×1010 CFU) on days -3 and 0, respectively, (n = 16; VB), or similarly inoculated with phosphate-buffered saline (n = 8; PBS). Venous and arterial blood were collected on -3, -1, 0, 2, 3, 5, 7, 10, and 14 relative to MH challenge for CBC, haptoglobin, and arterial blood oxygen saturation analysis. Continuously recorded variables included rumen temperature, activity, rumination, DMI, and feeding behavior. Data were analyzed with a repeated-measures mixed model (SAS 9.4) with fixed effects of day, inoculation, and the interaction. Bunk visit frequency and DMI were reduced (P &lt; 0.01) in VB steers throughout the 14 d post-MH period compared to PBS steers. Rumination (days 1, 7) and activity (days 1, 2, 4–8, and 11–13) were reduced (P &lt; 0.03) in VB steers vs PBS steers. Rumen temperature was elevated (P &lt; 0.04) in VB steers until day 6 post MH inoculation. Neutrophil concentrations (days 2 and 3), platelets (days 7–14), fibrinogen (days 2–10), and haptoglobin (days 2–7) were elevated (P &lt; 0.05) in VB vs PBS steers. Hematocrit was depressed (P &lt; 0.05) in VB steers on days 3–10. The VB steers had decreased (P &lt; 0.05) arterial SO2% and pO2 than PBS steers; however, the inoculation × day interaction was not significant. Results indicate that the experimental VB challenge substantially altered rumen temperature, DMI, feeding behavior, rumination and immunological response as expected. Arterial sO2% and pO2 concentrations were reduced minimally by VB challenge, indicating challenges in using blood gas to detect BRD in beef cattle.

Induction of protection in mice against a respiratory challenge by a vaccine formulated with exosomes isolated from Chlamydia muridarum infected cells

The goal of this study was to determine if exosomes, isolated from Chlamydia muridarum infected HeLa cells (C. muridarum-exosomes), induce protective immune responses in mice following vaccination using CpG plus Montanide as adjuvants. Exosomes, collected from uninfected HeLa cells and PBS, formulated with the same adjuvants, were used as negative controls. Mass spectrometry analyses identified 113 C. muridarum proteins in the C. muridarum-exosome preparation including the major outer membrane protein and the polymorphic membrane proteins. Vaccination with C. muridarum-exosomes elicited robust humoral and cell-mediated immune responses to C. muridarum elementary bodies. Following vaccination, mice were challenged intranasally with C. muridarum. Compared to the negative controls, mice immunized with C. muridarum-exosomes were significantly protected as measured by changes in body weight, lungs’ weight, and number of inclusion forming units recovered from lungs. This is the first report, of a vaccine formulated with Chlamydia exosomes, shown to elicit protection against a challenge.