ABSTRACT
Background & Aim: Ricin is one of the most lethal toxins, particularly if inhaled, and is considered a biological threat agent due to its wide availability and ease of production. Pulmonary ricin intoxication manifests in ARDS, cytokine storm, immune infiltration, and severe edema. Passive immunization is the preferred measure against pulmonary ricinosis, but only if administered shortly after exposure. Despite their potential to remedy pulmonary injury and inflammation, mesenchymal cell (MSC) therapies were never investigated in ricinosis. Here, we report the potential for treating pulmonary ricinosis with MesenCure, a professionalized allogeneic MSC therapy shown to reduce the mortality of patients suffering from severe pulmonary manifestations of COVID by 68%. Methods, Results & Conclusion(s): Preliminary studies demonstrated positive MesenCure effects in a sub-lethal pulmonary ricinosis model in CD1 mice. This model is regarded as highly translational due to the broad heterogeneity of these outbred mice. Positive effects included a reduction in excess protein content of the bronchoalveolar lavage fluid (BALF) by 45% when MesenCure was injected intravenously (IV) at 125k cells/animal, 48h post-exposure (PE) and evaluated one day later (p<0.05, Fig. 1A). Moreover, we found up to 52% reduction in the excess BALF leukocytes, when MesenCure was injected IV, 24h PE using the same dose (p<0.05, Fig. 1B) or 6h PE using a double dose (p<0.01, Fig. 1C), and evaluated two days PE. Optimizing the dose and administration route further improved the therapeutic outcome of MesenCure applied 6h PE as assessed by weight loss. As shown in Fig. 1D-E, IV injection of 250k-500k MesenCure cells/animal slightly protected the intoxicated animals against weight loss (p for treatment x time interaction <0.01 or <0.05 for 250k and 500k cells/animal, respectively). Interestingly, one million cells IV resulted in a lesser effect (not shown), however when injected subcutaneously (SC), 1M cells were very effective (p<0.001, Fig. 1F), seemingly even more effective than 2M cells/animal SC (Fig. 1G). Surprisingly, 2M thawed cells/animal injected SC protected the animals against weight loss almost completely (p<0.0001, Fig. H). In conclusion, we provide evidence for the potential of SC MSCs, specifically MesenCure, for treating pulmonary ricinosis and possibly other forms of ARDS. In agreement with Giri and Galipeau (2020), we provide further evidence for the dependency of MSC outcomes on their specific state and administration route. [Figure presented]Copyright © 2023 International Society for Cell & Gene Therapy
ABSTRACT
Methods: Here, we sought to investigate the effects of TLR7 pathway activation on mouse behaviour 24 hours post-activation. Female CD1 mice received an intraperitoneal injection of the synthetic TLR7 agonist, R848, or an equivalent volume of saline and were subjected to the Open Field and Forced Swim Test 24 hours later (n=10/group). Brain and liver tissues were then collected for downstream gene expression analysis. Result(s): Independent T-tests confirmed that systemic R848 challenge induced a strong peripheral and central inflammatory response, as indicated by a 250-fold increase in hepatic SAA-2 mRNA expression (p<0.0001) and a 75-fold increase in CXCL10 mRNA expression in the prefrontal cortex (p<0.01), relative to controls. These changes in inflammatory markers were accompanied by evidence of sickness behaviour - in particular, a decrease in exploratory rearing (p<0.01). Conclusion(s): This demonstrates that R848 can be used to create a model reflective of viral-like illness and provides a useful tool for investigating the behavioural effects of TLR7-mediated inflammation. To further these results, we aim to explore the metabolic consequences of LPS and R848 challenge and relate these to inflammatory and behavioural changes. This will accompany our data on the metabolic signatures of SARS-CoV-2-infected cells and animals, and of long-COVID patients. Copyright © 2022
ABSTRACT
Background: Currently available COVID-19 vaccination regimens in the US deliver either a homologous spike (S) mRNA prime-boost or a prime-only S DNA adenovirus-vectored antigen to elicit humoral and cell-mediated responses to confer protection against SAR-CoV-2 infection. Alternatively, heterologous vaccination using two different platforms has the potential to enhance and expand immune protection. Addition of a second SARS-CoV-2 antigen, the nucleocapsid (N) protein that is less subject to mutation and elicits vigorous T-cell responses, may also be advantageous. We report immunological responses to homologous and heterologous prime-boost vaccination regimens with a human DNA adenovirus serotype 5 S plus N (AdS+N) and/or a self-amplifying S-only mRNA vaccine (AAAH) delivered with a nanostructured lipid carrier (NLC). Methods: CD-1 mice received homologous or heterologous prime-boost combinations of AdS+N and AAAH. Priming doses were administered on Day 0, booster doses were delivered on Day 21, and mice were euthanized for blood and organ collection on Day 35. Serum was analyzed for anti-S (both wild type and variant) and anti-N IgG subtypes by ELISA. Spleen-resident CD4+ and CD8+ T cells were tested for IFN-γ, TNF-α, and IL-2 production in response to S-WT, S Delta variant and N protein overlapping peptides by intracellular cytokine staining (ICS). Splenocyte cytokine secretion upon stimulation with S-WT/N peptides was also assessed by IFN-γ and IL-4 ELISpot. Serum neutralization of the original Wuhan strain, Delta, and B.1.351 variants was assessed by a pseudovirus neutralization assay. Results: The highest humoral and T-cell responses were seen with the heterologous AAAH prime-AdS+N boost regimen, with a significant increase in T-cell responses relative to homologous vaccination. S protein-binding IgG was similar between wild type and Delta variant S proteins, with a strong/clear Th1/Th2 bias, and T cells responded to S wild type and S Delta peptides with similar levels of cytokine expression. Sera from AAAH prime-AdS+N boost mice showed the ability to neutralize Wuhan D614G, Delta, and B.1.351 (South Africa) variant pseudoviruses at high levels. Conclusion: Heterologous vaccination with the AAAH RNA vaccine prime and an AdS+N DNA boost may provide substantially improved humoral and cell-based immunity against SARS-CoV-2 variants by leveraging the advantages of each vaccine platform technology and by inclusion of immune responses to N.
ABSTRACT
Objectives: With over 90 million cases reported in the globe, the COVID-19 pandemic caused by SARS-CoV-2 has been a serious public health crisis. Development of novel and specific antiviral drugs against the SARS-CoV-2 has been an urgent demand. One such drug is Favipiravir, initially developed as an antiviral drug against influenza. Now Favipiravir has received approvals for emergency use against SARS-CoV-2 in many countries. A better understanding of Favipiravir’s biodistribution and pharmacokinetics in vivo will facilitate the clinical development of antiviral drugs against the SARS-CoV-2. Herein, we reported the evaluation of [18 F]Favipiravir with PET in cross-species studies to demonstrate the drug’s biodistribu-tion and pharmacokinetics and investigate the potentially increased risk of neurodegenerative diseases and/or neuroinflammation to COVID-19. Methods: The radiosynthesis of [18 F]Favipiravir was via labeling a commercially available precursor, methyl-5-chloroisoxazolo[4,5-b] pyrazine-3-carboxylate with K[18F]F/K222 and K2CO3 in DMSO at 130°C for 10 min, followed by hydrolysis with NaOH (aq.) at 110°C for 15 min.1 The whole body distribution on CD-1 mice was performed at four time points (5, 15, 30, 60 min). PET studies were carried out in CD-1 mice and AD mice (5XFAD) and naïve rhesus monkeys. We also performed the radiometabolite analysis of [18 F]Favipiravir in plasma and brain of CD-1 mice at 30 min post-injection. Results: [18 F]Favipiravir was obtained in 29% isolated radiochemi-cal yield (decay corrected). The radiochemical purity of the tracer was greater than 99%. No sign of radiolysis was observed for [18F] Favipiravir up to 120 min after formulation with 10% EtOH/saline. High radioactivity accumulation was observed in blood, lung, liver, kidney, and bone (around or more than 5% ID/g, injected dose per gram of wet tissue). The radioactivity level reached a plateau in small intestine, kidney and liver at 30,15 and 5 min, respectively, followed by slow washout, indicating that [18F]Favipiravir was possibly eliminated via the hepatobiliary and urinary pathway. For the radio-metabolic analysis of [18F]Favipiravir, average 41% and 89% of the radioactivity was parent fraction in the mice brain and plasma at 30 min post-injection (n=2), respectively. In PET imaging of CD-1 mice, the standard uptake value (SUV) of [18F]Favipiravir in brain reached its max value of 0.5 at 10 min and slowly reduced to 0.4 at 60 min. The results of PET imaging of AD mice with [18 F]Favipiravir were similar with that of CD-1 mice. In PET imaging of Rhesus monkeys, the brain uptake of [18 F]Favipiravir reached the max value of 0.5 SUV at 5 min and subsequently decreased to 50-60% of the maximum at 60 min. Conclusion: The evaluation of [18F]Favipiravir has demonstrated with bio-distribution and PET in mice and NHPs. Further evaluation of pharmacokinetics of [18F]Favipiravir in whole body monkey scans and LPS-induced neuroinflammation models is underway.