Highly stable and immunogenic CMV T cell vaccine candidate developed using a synthetic MVA platform.

Human cytomegalovirus (CMV) is the most common infectious cause of complications post-transplantation, while a CMV vaccine for transplant recipients has yet to be licensed. Triplex, a multiantigen Modified Vaccinia Ankara (MVA)-vectored CMV vaccine candidate based on the immunodominant antigens phosphoprotein 65 (pp65) and immediate-early 1 and 2 (IE1/2), is in an advanced stage of clinical development. However, its limited genetic and expression stability restricts its potential for large-scale production. Using a recently developed fully synthetic MVA (sMVA) platform, we developed a new generation Triplex vaccine candidate, T10-F10, with different sequence modifications for enhanced vaccine stability. T10-F10 demonstrated genetic and expression stability during extensive virus passaging. In addition, we show that T10-F10 confers comparable immunogenicity to the original Triplex vaccine to elicit antigen-specific T cell responses in HLA-transgenic mice. These results demonstrate improvements in translational vaccine properties of an sMVA-based CMV vaccine candidate designed as a therapeutic treatment for transplant recipients.

Functional SARS-CoV-2-specific T cells of donor origin in allogeneic stem cell transplant recipients of a T-cell-replete infusion: A prospective observational study.

In the current post-pandemic era, recipients of an allogeneic hematopoietic stem cell transplant (HCT) deserve special attention. In these vulnerable patients, vaccine effectiveness is reduced by post-transplant immune-suppressive therapy; consequently, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) disease (COVID-19) is often associated with elevated morbidity and mortality. Characterizing SARS-CoV-2 adaptive immunity transfer from immune donors to HCT recipients in the context of immunosuppression will help identify optimal timing and vaccination strategies that can provide adequate protection to HCT recipients against infection with evolving SARS-CoV-2 variants. We performed a prospective observational study (NCT04666025 at ClinicalTrials.gov) to longitudinally monitor the transfer of SARS-CoV-2-specific antiviral immunity from HCT donors, who were either vaccinated or had a history of COVID-19, to their recipients via T-cell replete graft. Levels, function, and quality of SARS-CoV-2-specific immune responses were longitudinally analyzed up to 6 months post-HCT in 14 matched unrelated donor/recipients and four haploidentical donor/recipient pairs. A markedly skewed donor-derived SARS-CoV-2 CD4 T-cell response was measurable in 15 (83%) recipients. It showed a polarized Th1 functional profile, with the prevalence of central memory phenotype subsets. SARS-CoV-2-specific IFN-γ was detectable throughout the observation period, including early post-transplant (day +30). Functionally experienced SARS-CoV-2 Th1-type T cells promptly expanded in two recipients at the time of post-HCT vaccination and in two others who were infected and survived post-transplant COVID-19 infection. Our data suggest that donor-derived SARS-CoV-2 T-cell responses are functional in immunosuppressed recipients and may play a critical role in post-HCT vaccine response and protection from the fatal disease. Clinical trial registration: clinicaltrials.gov, identifier NCT04666025.

Hematopoietic stem cell donor vaccination with cytomegalovirus triplex augments frequencies of functional and durable cytomegalovirus-specific T cells in the recipient: A novel strategy to limit antiviral prophylaxis.

To enhance protective cytomegalovirus (CMV)-specific T cells in immunosuppressed recipients of an allogeneic hematopoietic cell transplant (HCT), we evaluated post-HCT impact of vaccinating healthy HCT donors with Triplex. Triplex is a viral vectored recombinant vaccine expressing three immunodominant CMV antigens. The vector is modified vaccinia Ankara (MVA), an attenuated, non-replicating poxvirus derived from the vaccinia virus strain Ankara. It demonstrated tolerability and immunogenicity in healthy adults and HCT recipients, in whom it also reduced CMV reactivation. Here, we report feasibility, safety, and immunological outcomes of a pilot phase 1 trial (NCT03560752 at ClinicalTrials.gov) including 17 CMV-seropositive recipients who received an HCT from a matched related donor (MRD) vaccinated with 5.1 × 108 pfu/ml of Triplex before cell harvest (median 15, range 11-28 days). Donor and recipient pairs who committed to participation in the trial resulted in exceptional adherence to the protocol. Triplex was well-tolerated with limited adverse events in donors and recipients, who all engrafted with full donor chimerism. On day 28 post-HCT, levels of functional vaccinia- and CMV-specific CD137+ CD8+ T cells were significantly higher (p < .0001 and p = .0174, respectively) in recipients of Triplex vaccinated MRD than unvaccinated MRD (control cohort). Predominantly, central and effector memory CMV-specific T-cell responses continued to steadily expand through 1-year follow-up. CMV viremia requiring antivirals developed in three recipients (18%). In summary, this novel approach represents a promising strategy applicable to different HCT settings for limiting the use of antiviral prophylaxis, which can impair and delay CMV-specific immunity, leading to CMV reactivation requiring treatment.

Vaccine-induced spike- and nucleocapsid-specific cellular responses maintain potent cross-reactivity to SARS-CoV-2 Delta and Omicron variants.

Cell-mediated immunity may contribute to providing protection against SARS-CoV-2 and its variants of concern (VOC). We developed COH04S1, a synthetic multiantigen modified vaccinia Ankara (MVA)-based COVID-19 vaccine that stimulated potent spike (S) and nucleocapsid (N) antigen-specific humoral and cellular immunity in a phase 1 clinical trial in healthy adults. Here, we show that individuals vaccinated with COH04S1 or mRNA vaccine BNT162b2 maintain robust cross-reactive cellular immunity for six or more months post-vaccination. Although neutralizing antibodies induced in COH04S1- and BNT162b2-vaccinees showed reduced activity against Delta and Omicron variants compared to ancestral SARS-CoV-2, S-specific T cells elicited in both COH04S1- and BNT162b2-vaccinees and N-specific T cells elicited in COH04S1-vaccinees demonstrated potent and equivalent cross-reactivity against ancestral SARS-CoV-2 and the major VOC. These results suggest that vaccine-induced T cells to S and N antigens may constitute a critical second line of defense to provide long-term protection against SARS-CoV-2 VOC.

COH04S1 and beta sequence-modified vaccine protect hamsters from SARS-CoV-2 variants.

COVID-19 vaccine efficacy is threatened by emerging SARS-CoV-2 variants of concern (VOC) with the capacity to evade protective neutralizing antibody responses. We recently developed clinical vaccine candidate COH04S1, a synthetic modified vaccinia Ankara vector (sMVA) co-expressing spike and nucleocapsid antigens based on the Wuhan-Hu-1 reference strain that showed potent efficacy to protect against ancestral SARS-CoV-2 in Syrian hamsters and non-human primates and was safe and immunogenic in healthy volunteers. Here, we demonstrate that intramuscular immunization of Syrian hamsters with COH04S1 and an analogous Beta variant-adapted vaccine candidate (COH04S351) elicits potent cross-reactive antibody responses and protects against weight loss, lower respiratory tract infection, and lung pathology following challenge with major SARS-CoV-2 VOC, including Beta and the highly contagious Delta variant. These results demonstrate efficacy of COH04S1 and a variant-adapted vaccine analog to confer cross-protective immunity against SARS-CoV-2 and its emerging VOC, supporting clinical investigation of these sMVA-based COVID-19 vaccine candidates.

Safety and immunogenicity of a synthetic multiantigen modified vaccinia virus Ankara-based COVID-19 vaccine (COH04S1): an open-label and randomised, phase 1 trial.

Background: COH04S1, a synthetic attenuated modified vaccinia virus Ankara vector co-expressing SARS-CoV-2 spike and nucleocapsid antigens, was tested for safety and immunogenicity in healthy adults. Methods: This combined open-label and randomised, phase 1 trial was done at the City of Hope Comprehensive Cancer Center (Duarte, CA, USA). We included participants aged 18-54 years with a negative SARS-CoV-2 antibody and PCR test, normal haematology and chemistry panels, a normal electrocardiogram and troponin concentration, negative pregnancy test if female, body-mass index of 30 kg/m2 or less, and no modified vaccinia virus Ankara or poxvirus vaccine in the past 12 months. In the open-label cohort, 1·0 × 107 plaque-forming units (PFU; low dose), 1·0 × 108 PFU (medium dose), and 2·5 × 108 PFU (high dose) of COH04S1 were administered by intramuscular injection on day 0 and 28 to sentinel participants using a queue-based statistical design to limit risk. In a randomised dose expansion cohort, additional participants were randomly assigned (3:3:1), using block size of seven, to receive two placebo vaccines (placebo group), one low-dose COH04S1 and one placebo vaccine (low-dose COH04S1 plus placebo group), or two low-dose COH04S1 vaccines (low-dose COH04S1 group). The primary outcome was safety and tolerability, with secondary objectives assessing vaccine-specific immunogenicity. The primary immunological outcome was a four times increase (seroconversion) from baseline in spike-specific or nucleocapsid-specific IgG titres within 28 days of the last injection, and seroconversion rates were compared with participants who received placebo using Fisher's exact test. Additional secondary outcomes included assessment of viral neutralisation and cellular responses. This trial is registered with ClinicalTrials.gov, NCT046339466. Findings: Between Dec 13, 2020, and May 24, 2021, 56 participants initiated vaccination. On day 0 and 28, 17 participants received low-dose COH04S1, eight received medium-dose COH04S1, nine received high-dose COH04S1, five received placebo, 13 received low-dose COH04S1 followed by placebo, and four discontinued early. Grade 3 fever was observed in one participant who received low-dose COH04S1 and placebo, and grade 2 anxiety or fatigue was seen in one participant who received medium-dose COH04S1. No severe adverse events were reported. Seroconversion was observed in all 34 participants for spike protein and 32 (94%) for nucleocapsid protein (p<0·0001 vs placebo for each comparison). Four times or more increase in SARS-CoV-2 neutralising antibodies within 56 days was measured in nine of 17 participants in the low-dose COH04S1 group, all eight participants in the medium-dose COH04S1 group, and eight of nine participants in the high-dose COH04S1 group (p=0·0035 combined dose levels vs placebo). Post-prime and post-boost four times increase in spike-specific or nucleocapsid-specific T cells secreting interferon-γ was measured in 48 (98%; 95% CI 89-100) of 49 participants who received at least one dose of COH04S1 and provided a sample for immunological analysis. Interpretation: COH04S1 was well tolerated and induced spike-specific and nucleocapsid-specific antibody and T-cell responses. Future evaluation of this COVID-19 vaccine candidate as a primary or boost vaccination is warranted. Funding: The Carol Moss Foundation and City of Hope Integrated Drug Development Venture programme.

Synthetic multiantigen MVA vaccine COH04S1 protects against SARS-CoV-2 in Syrian hamsters and non-human primates.

Second-generation COVID-19 vaccines could contribute to establish protective immunity against SARS-CoV-2 and its emerging variants. We developed COH04S1, a synthetic multiantigen modified vaccinia Ankara-based SARS-CoV-2 vaccine that co-expresses spike and nucleocapsid antigens. Here, we report COH04S1 vaccine efficacy in animal models. We demonstrate that intramuscular or intranasal vaccination of Syrian hamsters with COH04S1 induces robust Th1-biased antigen-specific humoral immunity and cross-neutralizing antibodies (NAb) and protects against weight loss, lower respiratory tract infection, and lung injury following intranasal SARS-CoV-2 challenge. Moreover, we demonstrate that single-dose or two-dose vaccination of non-human primates with COH04S1 induces robust antigen-specific binding antibodies, NAb, and Th1-biased T cells, protects against both upper and lower respiratory tract infection following intranasal/intratracheal SARS-CoV-2 challenge, and triggers potent post-challenge anamnestic antiviral responses. These results demonstrate COH04S1-mediated vaccine protection in animal models through different vaccination routes and dose regimens, complementing ongoing investigation of this multiantigen SARS-CoV-2 vaccine in clinical trials.

Successful outcome of pre-engraftment COVID-19 in an HCT patient: impact of targeted therapies and cellular immunity.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has emerged as a global pandemic that upended existing protocols and practices, including those for allogeneic hematopoietic stem cell transplantation (HCT). Here, we describe the successful clinical course and multiple key interventions administered to an acute lymphoblastic leukemia patient, who tested SARS-CoV-2 positive by reverse transcriptase polymerase chain reaction on day -1 of matched unrelated donor (SARS-CoV-2 immunoglobulin G negative) T-cell-replete HCT. This experience allowed for implementing a virologic and immunomonitoring panel to characterize the impact of SARS-CoV-2 on the recipient's nascent humoral and cellular immune response. The finding of robust, functional, and persistent levels of SARS-CoV-2-specific T cells, starting early after transplant was unexpected, and in combination with the clinical strategy, may have contributed to the favorable outcome. Additionally, it is plausible that preexisting cross-reactive endemic coronavirus immunity in the allogeneic graft reduced recipient susceptibility to COVID-19 disease. This case supports the critical role that T-cell responses may play in mitigating SARS-CoV-2 infection, even in the context of transplant immunosuppression, in which reconstitution of humoral response is commonly delayed. Interventional approaches to transfer SARS-CoV-2-specific cellular immunity such as HCT donor vaccination and adaptive cellular therapy could be of benefit.

Synthetic Multiantigen MVA Vaccine COH04S1 Protects Against SARS-CoV-2 in Syrian Hamsters and Non-Human Primates.

Second-generation COVID-19 vaccines could contribute to establish protective immunity against SARS-CoV-2 and its emerging variants. We developed COH04S1, a synthetic multiantigen Modified Vaccinia Ankara-based SARS-CoV-2 vaccine that co-expresses spike and nucleocapsid antigens. Here, we report COH04S1 vaccine efficacy in animal models. We demonstrate that intramuscular or intranasal vaccination of Syrian hamsters with COH04S1 induces robust Th1-biased antigen-specific humoral immunity and cross-neutralizing antibodies (NAb) and protects against weight loss, lower respiratory tract infection, and lung injury following intranasal SARS-CoV-2 challenge. Moreover, we demonstrate that single-dose or two-dose vaccination of non-human primates with COH04S1 induces robust antigen-specific binding antibodies, NAb, and Th1-biased T cells, protects against both upper and lower respiratory tract infection following intranasal/intratracheal SARS-CoV-2 challenge, and triggers potent post-challenge anamnestic antiviral responses. These results demonstrate COH04S1-mediated vaccine protection in animal models through different vaccination routes and dose regimens, complementing ongoing investigation of this multiantigen SARS-CoV-2 vaccine in clinical trials.

BCG Cell Wall Skeleton As a Vaccine Adjuvant Protects Both Infant and Old-Aged Mice from Influenza Virus Infection.

Bacillus Calmette-Guerin (BCG) and the cell wall skeleton (CWS) derived from BCG are known to enhance nonspecific immune activation and anti-cancer immunity; however, their roles as a vaccine adjuvant are largely unknown. Here, we report that BCG-CWS acts as a strong immune adjuvant by promoting the protective immune responses in mouse models with influenza vaccination. The different aged mice immunized with inactivated split vaccine with or without BCG-CWS were challenged with an influenza pandemic virus. When protective immune responses were compared, even a single immunization of adult mice with a BCG-CWS-adjuvanted vaccine showed significantly enhanced humoral immune responses with increased IgG1 and IgG2a isotype antibodies. Importantly, the protective effects by the BCG-CWS adjuvant for influenza vaccination upon humoral and cellular immunogenicity were comparable between infants (6 days and 2 weeks old) and aged (20 months old) mice. Moreover, BCG-CWS dramatically augmented vaccine-mediated protective responses, including decreased viral loads, lung damage, and airway resistance, as well as increased mouse survival, amelioration of weight loss, and proinflammatory cytokine expression in all experimental groups including infant, adults, and old aged mice. We further provided the evidence that the BCG-CWS adjuvant effects were mediated through Toll-like receptors (TLR) 2 and TLR4 signaling pathways. Together, these data suggest that BCG-CWS can be promising as a potential influenza vaccine adjuvant in both young and old aged population through TLR2/4-mediated immune-boosting activities.

Natural killer cells contribute to enhanced respiratory disease after oil-in-water emulsion adjuvanted vaccination against respiratory syncytial virus and infection.

Respiratory syncytial virus (RSV) infection caused severe acute respiratory disease in children and the elderly. There is no licensed vaccine. It has been a challenging problem to avoid vaccine enhanced respiratory disease in developing a safe and effective RSV vaccine. Here, we investigated the impact of MF59-like oil-in-water emulsion adjuvant Addavax on the vaccine efficacy of inactivated split RSV (sRSV) and the roles of natural killer (NK) cells in enhanced respiratory disease in sRSV vaccinated mice after RSV infection. Addavax-adjuvanted sRSV vaccination induced higher levels of IgG1 isotype antibodies and more effective lung viral clearance upon RSV infection but promoted enhanced respiratory disease of weight loss, pulmonary inflammation, and NK and NK T (NKT) cell infiltrations in the lungs. Antibody treatment depleting NK cells prior to RSV infection resulted in preventing severe weight loss and histopathology, as well as attenuating infiltration of dendritic cell subsets and TNF-α+ T cells in the lungs. This study demonstrated the impacts of oil-in-water emulsion adjuvant on sRSV vaccination and the potential roles of NK and NKT cells in protection and respiratory disease after adjuvanted RSV vaccination and infection in a mouse model.

Development of a multi-antigenic SARS-CoV-2 vaccine candidate using a synthetic poxvirus platform.

Modified Vaccinia Ankara (MVA) is a highly attenuated poxvirus vector that is widely used to develop vaccines for infectious diseases and cancer. We demonstrate the construction of a vaccine platform based on a unique three-plasmid system to efficiently generate recombinant MVA vectors from chemically synthesized DNA. In response to the ongoing global pandemic caused by SARS coronavirus-2 (SARS-CoV-2), we use this vaccine platform to rapidly produce fully synthetic MVA (sMVA) vectors co-expressing SARS-CoV-2 spike and nucleocapsid antigens, two immunodominant antigens implicated in protective immunity. We show that mice immunized with these sMVA vectors develop robust SARS-CoV-2 antigen-specific humoral and cellular immune responses, including potent neutralizing antibodies. These results demonstrate the potential of a vaccine platform based on synthetic DNA to efficiently generate recombinant MVA vectors and to rapidly develop a multi-antigenic poxvirus-based SARS-CoV-2 vaccine candidate.

Natural Killer and CD8 T Cells Contribute to Protection by Formalin Inactivated Respiratory Syncytial Virus Vaccination under a CD4-Deficient Condition.

Respiratory syncytial virus (RSV) causes severe pulmonary disease in infants, young children, and the elderly. Formalin inactivated RSV (FI-RSV) vaccine trials failed due to vaccine enhanced respiratory disease, but the underlying immune mechanisms remain not fully understood. In this study, we have used wild type C57BL/6 and CD4 knockout (CD4KO) mouse models to better understand the roles of the CD4 T cells and cellular mechanisms responsible for enhanced respiratory disease after FI-RSV vaccination and RSV infection. Less eosinophil infiltration and lower pro-inflammatory cytokine production were observed in FI-RSV vaccinated CD4KO mice after RSV infection compared to FI-RSV vaccinated C57BL/6 mice. NK cells and cytokine-producing CD8 T cells were recruited at high levels in the airways of CD4KO mice, correlating with reduced respiratory disease. Depletion studies provided evidence that virus control was primarily mediated by NK cells whereas CD8 T cells contributed to IFN-γ production and less eosinophilic lung inflammation. This study demonstrated the differential roles of effector CD4 and CD8 T cells as well as NK cells, in networking with other inflammatory infiltrates in RSV disease in immune competent and CD4-deficient condition.