6 July 2021 Episode 150 : Mucosal immunity and vaccination: will intranasal vaccines be the “game changers” to prevent infection?

Episode 150 : Mucosal immunity and vaccination: will intranasal vaccines be the “game changers” to prevent infection?

Dear colleagues,

Several weeks ago one of you challenged me with the question “What about intranasal vaccination for SARS-COV-2”?  I was stimulated by the challenge, but then we went on holiday… However, as you see below, I took it up and try to answer the question in a systematic way, by first looking at the evidence that secretory IgA is important during infection and next explore ongoing studies with candidate intranasal vaccines.  

Please bear with me in this longest-ever, but very intriguing Episode.  

1. Importance of secretory IgA during COVID infections

Ep 150-1 : Russel argues about the importance of mucosal IgA:

• The nasopharynx associated lymphoid tissue (NALT) is known to be an inductive site for secretory IgA (SIgA) production in both the upper and the lower respiratory tract (URT and LRT).
• Bronchus-associated lymphoid tissue (BALT) is not normally present in adults but is found in children and adolescents, which may be one of the reasons why younger people are more resistant to COVID
• SIgA has  a number of effector functions, including prevention of adherence, and neutralization of viruses and even acts inside of epithelial cells. 
• While SIgA acts non-or anti-inflammatory, similar effector functions of IgG (dominant in the alveoli) are pro-inflammatory, hence could contribute to pathogenesis during infection.
• Intramuscular immunization (IM) mainly induces IgG, while intranasal (IN) immunization will induce SIgA in the URT and LRT.

Ep 150-2:  A logical deduction is that patients with serious primary immune deficiency (lacking B cells) or more common (and often asymptomatic) selective IgA deficiency could be more susceptible to COVID pathology.  The paper by Quinti et al shows that “the jury is still out”: there are indeed case reports that patients with primary immune deficiency have a protracted or recurrent course of the infection, but systematic studies are lacking.

Ep 150-3: Importance of secretory IgA in early and late response to SARS-CoV-2

• Early SARS-CoV-2–specific humoral responses were dominated by IgA antibodies
• IgA was more potent in virus neutralization than IgG
• While specific IgA  in serum decreased 1 month after the onset of symptoms, neutralizing IgA remained detectable in saliva for a longer time (days 49 to 73 post-symptoms)
• Also sIgA in broncho-alveolar lavage potently neutralizes SARS-CoV-2

Ep 150-4:  Nussenzweig’s group compared serum IgG and IgA (monomeric and dimeric) in a COVID convalescent cohort:

• Serum IgA neutralizing responses to SARS-CoV-2 correlate with IgG responses.
• The monomeric form of IgA found in serum is, twofold less potent than IgG,
• The dimeric, secretory form of IgA found in mucosa is 15 X more potent than their respective monomer forms against authentic SARS-CoV-2,

Suggesting that dimeric IgA is a more potent neutralizer than IgG

2. Reviews on mucosal immunity and vaccination

Ep 150-5: Park provides and overview of the immune response and existing SARS-CoV-2 vaccines and then argues in favor of intranasal administration

Ep 150-6: Tiboni provides another nice overview and describes in more detail the rationale and ongoing efforts for intranasal (IN) vaccination against SARS-CoV-2 pp. 12-17:

• Conceptually, it is evident that IM vaccination will mainly induce IgG and provide protection against lung disease after infection, leaving the possibility of replication of the virus in the upper airways, hence the risk for ongoing transmission.  On the contrary IN vaccination will induce more secretory IgA with the possibility to also protect upper airways against infection, hence “sterilizing immunity”.
• The intranasal route has also practical advantages:
  • No strict need of health care personnel or sterile environment
  • Better suited for mass vaccination, especially in children
  • Potentially lower dose will suffice
• Caveats:
  • Need for “bioadhesive” formulation: needs to remain sufficiently long to elicit immune responses
  • Potential allergic reactions with respiratory symptoms: e.g. egg-grown vaccines
  • Potential toxicity: e.g. inactivated viral subunit influenza vaccine (Nasalflu, Berna Biotech) was adjuvanted with E. coli heat-labile toxin and found after intranasal vaccination to increase the risk of Bell’s palsy.  Has been retracted
  • In case of SARS-CoV-2: risk of S-protein to enter the olfactory tissue t and potentially result in adverse effects (anosmia-aguesia or even neurological via lamina cribriformis)
• A lot of experimental IN vaccines for flu, RSV etc, but only one is on the market: cold-adapted live-attenuated influenza vaccines produced by Medimmune-AstraZeneca (Fluenz Tetra  in EU; NasoVac in India)  

3. Systematic discussion on some of the candidate IN SARS-CoV-2 vaccines

1. Chimp Adeno Ox1 vector:  Oxford/Jenner and Astra-Zeneca

Ep 150-7 A: is the Nature paper (July 2020) on the preclinical results with the Astra-Zeneca  Chimp Adeno Ox1 SARS-CoV-2 vaccine. After intramuscular (IM) vaccination and challenge of non-human primates they observed

• a significantly reduced viral load in the bronchoalveolar  lavage fluid and lower respiratory tract tissue and no pneumonia
• But no difference in nasal shedding between vaccinated and control macaques.

Ep  150-7 B: the same group in bioRxiv Jan 2021 on intranasal vaccination (IN) of rhesus macaques with the Astra-Zeneca construct resulted in reduced shedding and a reduction in viral load in bronchoalveolar lavage and lower respiratory tract tissue,  but no direct comparison with IM route.

In the same paper, Syrian hamsters were also vaccinated IM or IN and either challenged directly by instillation or via an infected cage mate (transmission).  Both experimental designs showed that IN was superior in protection against infection of oro-pharynx (but there was still virus present).

Importantly, in the direct challenge both IN and IM protected against lung infection and disease, but in the transmission experiment IN prevent completely against lung infection, while IM vaccination only partly.  

Ep 150-7 C: On the Clinical Trial.Gov site you find the outline of Phase I Study to Determine Safety, Tolerability and Immunogenicity of Intranasal Administration of the COVID Vaccine ChAdOx1 nCOV-19 in Healthy UK Adults.

2. Chimp Ad36 vector : University of Washington and Bharat Co

Ep 150-8 A: Hassan et all in Cell show very similar results in a direct comparison in mice: 

• Intramuscular Chimp-Adeno 36-SARS-CoV-2-S (prefusion as in the clinically used vaccines) induces robust systemic immune responses and protects against lung infection, inflammation, and pathology but does not confer sterilizing immunity, as evidenced by detection of viral RNA and induction of anti-nucleoprotein antibodies after SARS-CoV-2 challenge.
• Intranasal dose of ChAd-SARS-CoV-2-S induces high levels of neutralizing antibodies, promotes systemic and mucosal immunoglobulin A (IgA) and T cell responses, and almost entirely prevents SARS-CoV-2 infection in both the upper and lower respiratory tracts.

Ep 150-8 B: Based on this work, the Indian Bharat company (which produces the inactivated Covaxin) has submitted a phase1/2 protocol on the clinicaltrial.gov site

“ A Phase 1, Randomized, Double-blinded, Multicenter Study to Evaluate the Reactogenicity, Safety, and Immunogenicity of an Intranasal Adenovirus Vector COVID-19 Vaccine (BBV154) in Healthy Volunteers”

3. Human Ad 5 vector: Altimmune Inc

Ep 150-9 A: Feng in Nat. Comm shows that a single dose of Human Adeno-5 vector, expressing Spike , either IM or IN, can protect Chinese macaques against intratracheal challenge (although small viral blips were noted).

Ep 150-9 B: King et al use AdCOVID, an intranasal adenovirus type 5 (Ad5)-vectored vaccine encoding the receptor binding domain (RBD) of the SARS-CoV-2 spike protein, elicits a strong and focused immune response against RBD through the induction of mucosal IgA, serum neutralizing antibodies and CD4+ and CD8+ T cells with a Th1-like cytokine expression profile in mice.  

Ep 150-9 C: Disappointing press release: the company Altimmune, who is actively pursuing a similar intranasal vaccine for Influenza stops the phase ½ trial with AdCOVID, because  the magnitude of the response and the percent of subjects responding to AdCOVID were substantially lower than what had been demonstrated for other vaccines already authorized for emergency use.

4. Attenuated SARS-CoV-2 COVI-VAC: Codagenix

Ep 150-10 A: A very exciting PNAS on an attenuated SARS-CoV-2 in hamsters. 

• The attenuating is performed by codon pair de-optimization (= recoding a segment of the viral spike protein with synonymous suboptimal codon pairs) and deletion of the furin cleavage  site. 
• This virus still grows in vitro (but much less at 37 °C).  In hamsters growth is much less, there is no pathology, but it induces high neut Ab.
• It can be used as a very effective intranasal vaccine: Upon challenge with WT virus, COVI-VAC vaccination reduced lung challenge viral titers, resulted in undetectable virus in the brain, and protected hamsters from almost all SARS-CoV-2–associated weight loss.

Ep 150-10 B:    Press release announcing phase 1/3 trials with this candidate.

5. New Castle Disease vector: Lancaster University (UK)/ Biomedical research institute Texas (US).

Ep 150-11:  Park et al bioRxiv Jan 2021

• The recombinant (r)NDV-S vaccine expressing spike (S) protein of SARS-CoV-2 administrated via intranasal route in mice induced high levels of SARS-CoV-2-specific neutralizing immunoglobulin A (IgA) and IgG2a antibodies and T cell-mediated immunity.
• Hamsters vaccinated with two doses of vaccine showed complete protection from clinical disease including lung infection, inflammation, and pathological lesions after SARS-CoV2 challenge.
• Importantly, a single or double dose of intranasal rNDV-S vaccine completely blocked SARS-CoV-2 shedding in nasal turbinate and lungs within 4 days of vaccine administration in hamsters

6. Lentiviral vector: Institut Pasteur-TheraVectys Joint Laboratory

Ep 150-12: Ku et al show that an intraperitoneal prime, followed by intranasal boost with a lentiviral vector, expressing the VSV glycoprotein and the membrane-anchored SARS-CoV-2 Spike provide partial protection against infection and full protection against disease in mice and Syrian hamsters.

7. Vesiculo Stomatitis vector

Ep 150-13:   Interestingly, Furuyama et al found rapid protection from COVID-19 in nonhuman primates vaccinated intramuscularly but not intranasally with a single dose of a recombinant vaccine. 

• However, here they used no Adeno, but a licensed vesiculo-stomatitis EBOLA vaccine construct in which the SARS-CoV-2 spike was introduced (VSV-SARS2- EBOV).
• The IN vaccinated animals even showed enhanced COVID-19 pneumonia as compared to controls !!
• Fig 1 shows that neither the IM or IN vaccine protected against infection, but IM protected against disease.
• Neutralizing antibody titers were similar in IN and IM, but the cytolytic (Granzyme B+) CD8 T cell response after IM vaccination was much more robust in IM 

8. Respiratory Syncytium Virus vector: Meissa Vaccines

Ep 150-14 A: Meissa Vaccines (Kansas) announces a phase ½ trial using a SARS-CoV-2 Spike recombinant attenuated respiratory syncytium virus (RSV).

Ep 150-14 B:  The Nat Comm paper, describing this RSV vector, which has been attenuated by deletions and codon de-optimization.  This paper dates from 2016 and shows immonugenicity of this vector in cotton rats.

9. Subunit SARS-CoV-2 Spike vaccine with STING adjuvant: University of Houston

Ep 150-15: Xingue An bioRxiv July 2021

• A trimeric or monomeric S protein from the SARS-CoV-2 virus and using a liposomal, stimulator of interferon genes (STING), as an adjuvant.
• After a single intranasal administration of trimeric S-STING-liposones in Balb/c mice robust systemic B and T responses were observed, but also IgA responses in the BAL fluid and in the NALT and an increase in the number of total IgA secreting and S-specific IgA antibody secreting cells (ASCs).
• Very elaborated analysis; but neutralizing antibodies were only studied in serum

10. Subunit RBD with Salmonelle extracelluar vesicles:

Ep 150-16: A candidate based on extracellular vesicles (EVs) of Salmonella typhimurium that are decorated with the mammalian cell culture-derived Spike receptor-binding domain (RBD).  When used intranasally in hamsters, it was partly effective: avoided weight loss, had lower virus titers in bronchoalveolar lavage fluid, and experienced less severe lung pathology

Provisional Conclusions:

1. Studies in COVID patient provide clear evidence of the importance of potentially protective secretory IgA responses.

2. Recombinant Spike Adenoviral vectors for intranasal administration have provided good results in preclinical models, showing that they can induce neutralizing IgA and prevent infection and transmission. 
   1. Two Chimp Adeno’s have the strongest preclinical profile and are now in phase 1 trial.
   2. The preclinical results from human Ad5 were less convincing and a phase 1 trial has already been stopped for futility.    

3. Several other recombinant Spike viral vectors have been proposed as intranasal vaccines in preclinical studies.  Of these, the Newcastle Disease seems most promising, followed by a Lentiviral construct.  An RSV construct, of which I could not find SARS-CoV-2 preclinical data is also in phase 1. In contrast a  VSV Spike construct provided protection after IM administration, but enhanced disease after IN !   

4. Intriguingly an attenuated SARS-CoV-2 has passed the preclinical safety and efficacy thresholds and  is now in phase 1.

5. There are also two protein subunit vaccines in preclinical development for intranasal administration.

Obviously, phase 1 safety will be a major hurdle and olfactory/neurological side effects will have to be carefully monitored.  Disease enhancement should carefully be excluded.

In the future, it should be possible to test whether a IM prime, followed by IN boost could (or vice versa) could provide optimal protective immunity in upper and lower respiratory airways.  

I hope you are still with me in this fascinating journey…

Best wishes,