Co-administration of intranasal parainfluenza virus vaccines expressing antigenically distinct SARS-CoV-2 S antigens elicits broad and durable immunity in hamsters

ABSTRACT Intranasal COVID-19 vaccines with the ability to induce broad and durable mucosal and systemic immunity would be useful as stand-alone vaccines or in combination with injectable vaccines. We evaluated in hamsters the breadth of immunity and durability of protection elicited by co-administration of two live-attenuated bovine/human parainfluenza virus type 3 (B/HPIV3) vectors expressing antigenically distinct prefusion-stabilized S proteins of the ancestral SARS-CoV-2 isolate (B/HPIV3/S-6P) or the BA.5/Omicron variant (B/HPIV3/S-BA.5-2P). These vectors are being developed as bivalent pediatric vaccines against HPIV3 and SARS-CoV-2 and are based on bovine PIV3 with the fusion (F) and hemagglutinin-neuraminidase (HN) glycoproteins replaced by those of human PIV3. To broaden the S-specific antibody response, we evaluated co-administration of these B/HPIV3 S-expressing vectors. Each B/HPIV3 S-expressing vector induced robust serum anti-S IgG and IgA antibody levels to the antigen-matched S protein that were sustained for at least 5 months. Co-administration increased the breadth of the S-specific antibody response, spanning the breadth of the response elicited by each vector individually. Hamsters immunized intranasally with the mixture of vectors developed neutralizing antibodies to ancestral and recently circulating SARS-CoV-2 strains and were protected against BA.5/Omicron challenge 5 months after immunization, with no weight loss, SARS-CoV-2 challenge virus replication, or increase in host inflammatory cytokines in the airways after the challenge, indicating durable protection. Thus, intranasal co-administration of live-attenuated B/HPIV3 expressing antigenically distinct S proteins induced broad and durable antibody responses, and long-term protection against BA.5/Omicron challenge. This approach warrants further development and may better protect against emerging SARS-CoV-2 variants. IMPORTANCE Intranasal SARS-CoV-2 vaccines that induce systemic and local mucosal immunity hold promise, especially as booster vaccines against SARS-CoV-2 variants. We developed live-attenuated bovine/human parainfluenza virus type 3 (B/HPIV3)-vectored vaccines as bivalent pediatric vaccines against PIV3 and SARS-CoV-2. Co-administration in a single dose of two B/HPIV3 vectors, one expressing the spike protein S from the ancestral Wuhan-Hu-1 strain and one expressing S of BA.5/Omicron, induced broad serum anti-S antibody responses that remained strong for at least 5 months. Hamsters challenged with the BA.5/Omicron strain 5 months after immunization were protected from weight loss, inflammatory responses, and challenge virus replication in the airways. Thus, in the hamster model, intranasal co-administration of live-attenuated B/HPIV3 expressing SARS-CoV-2 S elicits protection durable over several months, and combining two antigenically distinct B/HPIV3 S-expressing vectors substantially broadens the antibody response. This mucosal immunization approach may provide durable and broad protection against SARS-CoV-2.