Dose sparing effects of novel adjuvants and aluminium hydroxide on two different vaccines in a neonatal mouse model

Jenny Lorena Molina Estupiñan^1,2Poorya Foroutan Pajoohian^1,2Gabriel Pedersen³Dennis Christensen³Serena Marchi⁴Emanuele Montomoli^4,5Stefanía P. Bjarnarson^1,2Ingileif Jonsdottir^1,2Auður Anna Aradottir Pind^1,2*

• ¹University of Iceland, Reykjavik, Iceland
• ²Landspitali, the National University Hospital of Iceland, Reykjavík, Iceland
• ³Center for Vaccine Research, Statens Serum Institut, Copenhagen, Denmark
• ⁴Department of Molecular and Developmental Medicine, University of Siena, Siena, Italy
• ⁵VisMederi srl, Siena, Italy

Childhood vaccination provides protection against infectious diseases, but multiple vaccinations are required to achieve this. In situations like influenza epidemics or COVID-19 pandemic, vaccine demands may exceed production capacity, highlighting the need for dose sparing strategies. Adjuvants can boost and modulate immune responses to vaccines and could reduce antigen doses needed to confer protection. Herein, we evaluated the dose sparing effects of the novel adjuvants dmLT, mmCT, CAF01 and CAF08b and alum (aluminium hydroxide) on primary neonatal antibody (Ab) response to a conjugate vaccine against Streptococcus pneumoniae, Pn1-CRM197, and a recombinant influenza hemagglutinin (HA) protein vaccine. Primary Ab levels of neonatal mice immunized once with a full dose of Pn1-CRM197 or HA were low. mmCT and CAF08b enhanced Pn1-specific IgG Abs elicited by fractional doses of Pn1-CRM197 providing 8-fold dose sparing of the vaccine, whereas dmLT and CAF01 provided 5-fold and 2-fold dose sparing, respectively. These adjuvants elicited protective Pn1-specific Ab levels against bacteremia (91%-63%) and pneumonia (50%-38%) in neonatal mice, when combined with half-dose of Pn1-CRM197. In addition, mmCT, CAF01 and CAF08b enhanced the persistence of Pn1-specific IgG Ab-secreting cells (ASCs) in bone marrow compared with full dose of vaccine only. With the influenza HA vaccine, CAF08b provided 40-fold dose sparing while CAF01 and mmCT provided 2-fold dose sparing. CAF08b induced micro-neutralization (MN) titers above protective levels in 100% and 86% of mice receiving 1/8th and 1/40th of HA dose, respectively, and CAF01 in 88% and 50% of the mice receiving 1/4th and 1/8th dose of HA, respectively, whereas only 38% of mice receiving full dose HA without adjuvant reached protective MN levels. Furthermore, these adjuvants provided cross-protective Abs and ASCs against a closely related heterologous influenza strain. In contrast, aluminium hydroxide did not provide any dose sparing effects. Collectively, our results demonstrate that mmCT, CAF01 and CAF08b enhanced protective humoral responses and had large dose sparing effects on both Pn1-CRM197 and HA vaccines, although the adjuvant effect was clearly vaccine-dependent. The results support potential use of safe adjuvants in situations when vaccine production capacity is limited, including vaccination of pediatric populations that may be of high risk.