Combination Ad26.RSV.preF/preF protein vaccine induces superior protective immunity compared with individual vaccine components in preclinical models

www.nature.com/npjvaccines ARTICLE OPEN Combination Ad26.RSV.preF/preF protein vaccine induces superior protective immunity compared with individual vaccine components in preclinical models 1234567890():,; Eirikur Saeland 1 ✉, Leslie van der Fits1, Renske Bolder 1, Marjolein Heemskerk-van der Meer1, Joke Drijver1, Yolinda van Polanen Cornelis Vaneman1, Lisanne Tettero1, Freek Cox1, Jan Serroyen1, Matthew J. Jorgensen 2, Johannes P. M. Langedijk 1, Hanneke Schuitemaker 1, Benoit Callendret1 and Roland C. Zahn 1 1 , Respiratory syncytial virus (RSV) is a leading cause of severe respiratory disease for which no licensed vaccine is available. We have previously shown that a prefusion (preF) conformation-stabilized RSV F protein antigen and an adenoviral vector encoding RSV preF protein (Ad26.RSV.preF) are immunogenic and protective in animals when administered as single components. Here, we evaluated a combination of the 2 components, administered as a single injection. Strong induction of both humoral and cellular responses was shown in RSV-naïve and pre-exposed mice and pre-exposed African green monkeys (AGMs). Both components of the combination vaccine contributed to humoral immune responses, while the Ad26.RSV.preF component was the main contributor to cellular immune responses in both mice and AGMs. Immunization with the combination elicited superior protection against RSV A2 challenge in cotton rats. These results demonstrate the advantage of a combination vaccine and support further clinical development. npj Vaccines (2023)8:45 ; https://doi.org/10.1038/s41541-023-00637-7 INTRODUCTION Respiratory syncytial virus (RSV) infections are a common cause of severe respiratory disease in older and immunocompromised adults1, children2, and those with certain chronic cardiac or pulmonary comorbidities3. Annually, RSV is estimated to cause >60 million acute respiratory infections (ARIs) in adults and children worldwide4. A recent prospective global study found that RSV causes disease burden in infected adults similar to or worse than influenza, as measured by clinical symptom scores and medical resource utilization5. Older adults are at particularly high risk for severe RSV-mediated disease; in the United States alone, there are an estimated 177,000 hospitalizations and 14,000 deaths due to RSV in adults aged ≥65 years6. The high RSV- associated morbidity in older adults may result from waning or impaired immune responses with age, referred to as “immune senescence”7,8. Given the high morbidity and significant disease burden of RSV in older adults, an effective prophylactic vaccine is needed to protect this population. The RSV fusion glycoprotein (RSV F) is an attractive vaccine antigen because it is the main target of RSV virus-neutralizing antibodies (VNAs) in human sera9,10. RSV F fuses the viral and host cell membranes by irreversible protein refolding from the labile prefusion (preF) conformation to the stable postfusion (postF) conformation11. RSV postF-based vaccines have shown limited or no protective efficacy in clinical trials12, possibly because human RSV virus-neutralizing antibodies (VNAs) are primarily formed against RSV preF protein10,13. RSV preF-based vaccine development has long been hindered by the instability of RSV preF; however, recent efforts to stabilize the protein in the preF conformation using mutations identified by structure-based design have been successful11,14. Compared with postF-based vaccines, RSV preF-based vaccines have demonstrated superior efficacy and immunogenicity in preclinical studies11,14–16. Although there is no commonly agreed-upon correlate of protection (CoP) against RSV infection, there is a wealth of literature establishing the importance of VNAs for preventing RSV infection17. VNAs and their correlation with protection are typically measured in the serum; however, some evidence exists that mucosal RSV-specific immunoglobulins (Igs), particularly IgA, are significantly associated with reduced RSV infection rates and disease severity18,19. Prior studies also suggest that CD4+ and CD8+ T cells play an important role in the immune response to RSV infection17,20. Airway CD8+ T cells have been shown to play a protective role against RSV disease in humans21 and mice22,23, whereas CD4+ follicular helper T cells support antibody production, class switching, and memory B-cell formation24. An effective RSV vaccine will preferably induce robust humoral (VNA titers [VNT]) and cellular (CD4+ and CD8+ T-cell) responses25,26. Subunit vaccines based on pre-fusion conformation-stabilized RSV F protein (RSV preF protein) have shown robust induction of RSV VNT in preclinical models27,28, and phase 1 and 2 clinical trials have demonstrated their immunogenicity and tolerability29,30. Ad26.RSV.preF is a recombinant, replication-incompetent, adenovirus type 26 (Ad26)-vector encoding a conformation-stabilized RSV preF protein that effectively induces both humoral and cellular responses in animal models15,27. Ad26.RSV.preF has been evaluated for safety and immunogenicity in clinical trials; a single dose of Ad26.RSV.preF was safe and induced lasting immune responses in older adults (age ≥60 years)31 and demonstrated protection against RSV in a recent human challenge study of adults (ages 18–50 years)32. 1 Janssen Vaccines & Prevention B.V., Leiden, The Netherlands. 2Department of Pathology, Section on Comparative Medicine, Wake Forest School of Medicine, Winston-Salem, NC, USA. ✉email: Published in partnership with the Sealy Institute for Vaccine Sciences E. Saeland et al. 2 1234567890():,; The combined data from nonclinical and clinical studies indicate that while Ad26.RSV.preF induces robust humoral and cellular immune responses, RSV preF protein subunit vaccines appear to induce superior humoral immune responses compared with Ad26.RSV.preF27. To date, no published work has investigated the advantages of combining an RSV protein subunit vaccine with a vector-based vaccine. Here, we assessed the immunogenicity and efficacy of a co-administered combination of Ad26.RSV.preF and RSV preF protein in a single injection and compared these with the responses to the individual components. RESULTS RSV preF protein significantly contributes to humoral immune responses, whereas Ad26.RSV.preF strongly induces cellular responses when administered as a combination in naïve mice To evaluate the potential advantages of combining Ad26.RSV.preF and RSV preF protein in a single vaccine, different dose levels of RSV preF protein (0.015 µg, 0.15 µg, 1.5 µg, and 15 µg) were combined with a low dose (1 × 108 viral particles [vp]) of Ad26.RSV.preF and immunogenicity was compared with Ad26.RSV.preF alone in RSV-naïve mice. Additionally, a combination of high-dose Ad26.RSV.preF (1 × 109 vp) and high-dose RSV preF protein (15 µg) was evaluated. Prime-boost immunization with combination regimens at Weeks 0 and 4 induced strong humoral and cellular immune responses mea (...truncated)