Toward a pan-SARS-CoV-2 vaccine targeting conserved epitopes on spike and non-spike proteins for potent, broad and durable immune responses.

BACKGROUND: The SARS-CoV-2 non-Spike (S) structural protein targets on nucleocapsid (N), membrane (M) and envelope (E), critical in the host cell interferon response and memory T-cell immunity, are grossly overlooked in COVID vaccine development. The current Spike-only vaccines bear an intrinsic shortfall for promotion of a fuller T cell immunity. Vaccines designed to target conserved epitopes could elicit strong cellular immune responses that would synergize with B cell responses and lead to long-term vaccine success. We pursue a universal (pan-SARS-CoV-2) vaccine against Delta, Omicrons and ever-emergent new mutants. METHODS AND FINDINGS: We explored booster immunogenicity of UB-612, a multitope-vaccine that contains S1-RBD-sFc protein and sequence-conserved promiscuous Th and CTL epitope peptides on the Sarbecovirus N, M and S2 proteins. To a subpopulation (N = 1,478) of infection-free participants (aged 18-85 years) involved in a two-dose Phase-2 trial, a UB-612 booster (third dose) was administered 6-8 months after the second dose. The immunogenicity was evaluated at 14 days post-booster with overall safety monitored until the end of study. The booster induced high viral-neutralizing antibodies against live Wuhan WT (VNT50, 1,711) and Delta (VNT50, 1,282); and against pseudovirus WT (pVNT50, 11,167) vs. Omicron BA.1/BA.2/BA.5 variants (pVNT50, 2,314/1,890/854), respectively. The lower primary neutralizing antibodies in the elderly were uplifted upon boosting to approximately the same high level in young adults. UB-612 also induced potent, durable Th1-oriented (IFN-γ+-) responses (peak/pre-boost/post-boost SFU/106 PBMCs, 374/261/444) along with robust presence of cytotoxic CD8+ T cells (peak/pre-boost/post-boost CD107a+-Granzyme B+, 3.6%/1.8%/1.8%). This UB-612 booster vaccination is safe and well tolerated without SAEs. CONCLUSIONS: By targeting conserved epitopes on viral S2, M and N proteins, UB-612 could provide potent, broad and long-lasting B-cell and T-cell memory immunity and offers the potential as a universal vaccine to fend off Omicrons and new VoCs without resorting to Omicron-specific immunogens. TRIAL REGISTRATION: ClinicalTrials.gov ID: NCT04773067; ClinicalTrials.gov ID: NCT05293665; ClinicalTrials.gov ID: NCT05541861.

A novel RBD-protein/peptide vaccine elicits broadly neutralizing antibodies and protects mice and macaques against SARS-CoV-2.

The development of safe and effective vaccines to respond to COVID-19 pandemic/endemic remains a priority. We developed a novel subunit protein-peptide COVID-19 vaccine candidate (UB-612) composed of: (i) receptor binding domain of SARS-CoV-2 spike protein fused to a modified single-chain human IgG1 Fc; (ii) five synthetic peptides incorporating conserved helper and cytotoxic T lymphocyte (Th/CTL) epitopes derived from SARS-CoV-2 structural proteins (three from S2 subunit, one from membrane and one from nucleocapsid), and one universal Th peptide; (iii) aluminum phosphate as adjuvant. The immunogenicity and protective immunity induced by UB-612 vaccine were evaluated in four animal models: Sprague-Dawley rats, AAV-hACE2 transduced BALB/c mice, rhesus and cynomolgus macaques. UB-612 vaccine induced high levels of neutralizing antibody and T-cell responses, in all animals. The immune sera from vaccinated animals neutralized the SARS-CoV-2 original wild-type strains and multiple variants of concern, including Delta and Omicron. The vaccination significantly reduced viral loads, lung pathology scores, and disease progression after intranasal and intratracheal challenge with SARS-CoV-2 in mice, rhesus and cynomolgus macaques. UB-612 has been tested in primary regimens in Phase 1 and Phase 2 clinical studies and is currently being evaluated in a global pivotal Phase 3 clinical study as a single dose heterologous booster.

A multitope SARS-CoV-2 vaccine provides long-lasting B cell and T cell immunity against Delta and Omicron variants.

BackgroundThe Delta and Omicron variants of SARS-CoV-2 are currently responsible for breakthrough infections due to waning immunity. We report phase I/II trial results of UB-612, a multitope subunit vaccine containing S1-RBD-sFc protein and rationally designed promiscuous peptides representing sarbecovirus conserved helper T cell and cytotoxic T lymphocyte epitopes on the nucleocapsid (N), membrane (M), and spike (S2) proteins.MethodWe conducted a phase I primary 2-dose (28 days apart) trial of 10, 30, or 100 µg UB-612 in 60 healthy young adults 20 to 55 years old, and 50 of them were boosted with 100 µg of UB-612 approximately 7 to 9 months after the second dose. A separate placebo-controlled and randomized phase II study was conducted with 2 doses of 100 µg of UB-612 (n = 3,875, 18-85 years old). We evaluated interim safety and immunogenicity of phase I until 14 days after the third (booster) dose and of phase II until 28 days after the second dose.ResultsNo vaccine-related serious adverse events were recorded. The most common solicited adverse events were injection site pain and fatigue, mostly mild and transient. In both trials, UB-612 elicited respective neutralizing antibody titers similar to a panel of human convalescent sera. The most striking findings were long-lasting virus-neutralizing antibodies and broad T cell immunity against SARS-CoV-2 variants of concern (VoCs), including Delta and Omicron, and a strong booster-recalled memory immunity with high cross-reactive neutralizing titers against the Delta and Omicron VoCs.ConclusionUB-612 has presented a favorable safety profile, potent booster effect against VoCs, and long-lasting B and broad T cell immunity that warrants further development for both primary immunization and heterologous boosting of other COVID-19 vaccines.Trial RegistrationClinicalTrials.gov: NCT04545749, NCT04773067, and NCT04967742.FundingUBI Asia, Vaxxinity Inc., and Taiwan Centers for Disease Control, Ministry of Health and Welfare.

[Ala499Val (C > T) and Lys939Gln (A > C) polymorphisms of the XPC gene: their correlation with male infertility].

OBJECTIVE: The Ala499Val (C > T) and Lys939Gln (A > C) of the XPC gene are two potentially functional nonsynonymous polymorphisms, which affect the rate of DNA repair and might change XPC production and activity. This study aimed to explore the distribution of these two polymorphisms in the Chinese Han population and their relationship with male infertility. METHODS: We genotyped the two polymorphisms of the XPC gene by the PCR-restriction fragment length polymorphism (PCR-RFLP) method in 318 infertile patients and 228 fertile male controls, detected the frequency of the alleles, and analyzed both the individual and the joint contribution of the two polymorphisms to male infertility. RESULTS: For the Ala499Val (C > T) polymorphism, the frequencies of the CC, CT, and TT genotypes were significantly different in distribution between the patients and the controls (P = 0.020). Males with the TT genotype had a lower risk of male infertility than those with the CC genotype (adjusted OR = 0.49, 95% CI: 0.23-0.88), and even lower than those with both CC and CT genotypes (adjusted OR = 0.39, 95% CI: 0.22-0.71). The Lys939Gln (A > C) polymorphism was not related with male infertility. The combined genotype analysis showed that the individuals with 1-4 risk alleles had a significantly higher risk of male infertility (adjusted OR = 2.75, 95% CI = 1.50-5.04) than those with 0 risk allele. CONCLUSION: The Ala499Val (C > T) polymorphism of the XPC gene is correlated with male infertility and may be a potential genetic risk factor for male infertility in the Chinese Han population.

Non-FK506-binding protein-12 neuroimmunophilin ligands increase neurite elongation and accelerate nerve regeneration.

Neurotrophic activity of neuroimmunophilin ligands (FK506 and its nonimmunosuppressant derivatives) has been assumed to be mediated by the FK506-binding protein-12 (FKBP-12). We recently showed that activity is retained in hippocampal neurons from FKBP-12 knockout mice, indicating that binding to FKBP-12 is not necessary. Here we show that three nonimmunosuppressant FK506 derivatives (V-13,450, V-13,629, and V-13,670) that do not bind FKBP-12 (>12.5 mM affinity) are equipotent to FKBP-12 ligands (FK506, V-10,367, and V-13,449) for increasing neurite elongation in SH-SY5Y cells. One non-FKBP-12 ligand (V-13,670) is also shown to accelerate functional recovery and nerve regeneration in the rat sciatic nerve crush model. Surprisingly, it exhibited an unusual dose-response effect upon oral administration, showing a novel bimodal dose-response for behavioral functional recovery and myelination, but not for axonal size, suggesting both Schwann cell and neuronal targets. Orally active non-FKBP-12 neuroimmunophilin ligands may be useful for the treatment of human neurological disorders without any potential side effects resulting from FKBP-12 binding.

Calpain inhibition protects against Taxol-induced sensory neuropathy.

Taxol is a highly effective anticancer agent that causes peripheral neuropathy as its major toxic side effect. The neuropathy is characterized by degeneration of sensory axons that may be severe enough to be dose limiting. Axonal degeneration involves the activation of the calcium-activated proteases calpains, and here we tested whether systemic inhibition of calpains with the peptide alpha-ketoamide calpain inhibitor AK295 can reduce the clinical and pathological effects of Taxol in a rodent model of Taxol neuropathy. In mice with Taxol neuropathy, AK295 reduced the degree of axonal degeneration in sensory nerve roots, and improved clinical measures of neuropathy, including behavioural and electrophysiological function. These findings were consistent for both 3- and 6-week models of neuropathy. In vitro, Taxol caused activation of both calpains and caspases in PC12 cells. AK295 inhibited the activation of calpains but did not interfere with the antimitotic effects of Taxol on microtubules, nor did it inhibit caspase-mediated cell death. These data implicate calpains in the pathogenesis of Taxol neuropathy, and demonstrate that AK295 can prevent axonal degeneration and clinical neuropathy in mice. In addition, AK295 did not interfere with the primary antineoplastic effects of Taxol on microtubules and cell death, suggesting that systemic calpain inhibition may be a good strategy for preventing neuropathy in patients being treated with Taxol.

WldS mice are resistant to paclitaxel (taxol) neuropathy.

The WldS mouse is a unique mutant strain that demonstrates the remarkable phenotype of prolonged survival of transected axons ("slow Wallerian degeneration"). In these studies, we tested whether this neuroprotective phenotype extends to axonal degeneration seen in a progressive peripheral neuropathy. WldS and wild-type mice were intoxicated with the cancer chemotherapeutic agent paclitaxel (Taxol). The severity of the resultant sensory neuropathy was compared with behavioral, physiological, and pathological measures. WldS mice were resistant to paclitaxel neuropathy by all measures, and the resistance was because of protection against axonal degeneration. These studies demonstrate for the first time that the WldS mouse is more than a slow Wallerian degeneration phenotype, emphasizing the mechanistic link between Wallerian degeneration and peripheral neuropathy. Understanding how this mutant gene confers protection against axonal degeneration will provide important clues toward prevention of axonal degeneration in several human neurological disorders.

[Clinical application and experimental studies of the pulsed inhaled nitric oxide flow controlling instrument].

A flow controlling system for pulsed inhaled nitric oxide has been developed and tested, and here its features and initial animal experiments and clinical applications are described. The physical characteristic test indicates that the practical released dose of NO gas is very close to the theoretical flow of NO gas at variant pressures. Animal experiments demonstrate that inhaled NO gas concentration is lower than the concentration of theoretical inhalation, but the variance is not remarkable (p>0.05). When sixteen cases with CHD and PH were chosen to inhale NO gas (15 ppm, 15 min) PAP and PVR of all cases were reduced after inhalation of NO gas from 617 +/-51.3 dyn x s x cm(-5), 54.4+/-13.1 mmHg to 417+/-36.9 dym x s x cm(-5), 33.8+/-12.3 mmHg (PVR, p<0.01; PAP, p<0.01) respectively. When gas inhalation was stopped, these values returned to their base lines after a short period of time. All these show that the pulsed inhaled NO flow controlling instrument in accordance with the requirements of the designing, can be widely used in clinical diagnoses and treatments and will be a new tool offered for the treatments of the patients with PH.