COVID-19 is a novel disease caused by SARS-CoV-2 which has conquered the world rapidly resulting in a pandemic that massively impacts our health, social activities, and economy. It is likely that vaccination is the only way to form “herd immunity” and restore the world to normal. Here we developed a vaccine candidate for COVID-19 based on the virus-like particle AP205 displaying the spike receptor binding motif (RBM), which is the major target of neutralizing antibodies in convalescent patients. To this end, we genetically fused the RBM domain of SARS-CoV-2 to the C terminus of AP205 of dimerized capsid proteins. The fused VLPs were expressed in E. coli, which resulted in insoluble aggregates. These aggregates were denatured in 8 M urea followed by refolding, which reconstituted VLP formation as confirmed by electron microscopy analysis. Importantly, immunized mice were able to generate high levels of IgG antibodies recognizing eukaryotically expressed receptor binding domain (RBD) as well as spike protein of SARS-CoV-2. Furthermore, induced antibodies were able to neutralize SARS-CoV-2/ABS/NL20. Additionally, this vaccine candidate has the potential to be produced at large scale for immunization programs.

Background: SARS‐CoV‐2 caused one of the most devastating pandemics in the recent history of mankind. Due to various countermeasures, including lock‐downs, wearing masks, and increased hygiene, the virus has been controlled in some parts of the world. More recently, the availability of vaccines, based on RNA or adenoviruses, has greatly added to our ability to keep the virus at bay; again, however, in some parts of the world only. While available vaccines are effective, it would be desirable to also have more classical vaccines at hand for the future. Key feature of vaccines for long‐term control of SARS‐CoV‐2 would be inexpensive production at large scale, ability to make multiple booster injections, and long‐term stability at 4℃.Methods: Here, we describe such a vaccine candidate, consisting of the SARS‐CoV‐2 receptor‐binding motif (RBM) grafted genetically onto the surface of the immunologically optimized cucumber mosaic virus, called CuMV_TT‐RBM.Results: Using bacterial fermentation and continuous flow centrifugation for purification, the yield of the production process is estimated to be >2.5 million doses per 1000‐litre fermenter run. We demonstrate that the candidate vaccine is highly immunogenic in mice and rabbits and induces more high avidity antibodies compared to convalescent human sera. The induced antibodies are more cross‐reactive to mutant RBDs of variants of concern (VoC). Furthermore, antibody responses are neutralizing and long‐lived. In addition, the vaccine candidate was stable for at least 14 months at 4℃.Conclusion: Thus, the here presented VLP‐based vaccine may be a good candidate for use as conventional vaccine in the long term.

The ongoing coronavirus disease (COVID-19) pandemic is caused by a new coronavirus (severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2)) first reported in Wuhan City, China. From there, it has been rapidly spreading to many cities inside and outside China. Nowadays, more than 110 million cases with deaths surpassing 2 million have been recorded worldwide, thus representing a major health and economic issues. Rapid development of a protective vaccine against COVID-19 is therefore of paramount importance. Here, we demonstrated that the recombinantly expressed receptor-binding domain (RBD) of the spike protein can be coupled to immunologically optimized virus-like particles derived from cucumber mosaic virus (CuMV_TT). The RBD displayed CuMV_TT bound to ACE2, the viral receptor, demonstrating proper folding of RBD. Furthermore, a highly repetitive display of the RBD on CuMV_TT resulted in a vaccine candidate that induced high levels of specific antibodies in mice, which were able to block binding of the spike protein to ACE2 and potently neutralize SARS-CoV-2 virus in vitro.

Background: The highly contagious SARS‐CoV‐2 is mainly transmitted by respiratory droplets and aerosols. Consequently, people are required to wear masks and maintain a social distance to avoid spreading of the virus. Despite the success of the commercially available vaccines, the virus is still uncontained globally. Given the tropism of SARS‐CoV‐2, a mucosal immune reaction would help to reduce viral shedding and transmission locally. Only seven out of hundreds of ongoing clinical trials are testing the intranasal delivery of a vaccine against COVID‐19.Methods: In the current study, we evaluated the immunogenicity of a traditional vaccine platform based on virus‐like particles (VLPs) displaying RBD of SARS‐CoV‐2 for intranasal administration in a murine model. The candidate vaccine platform, CuMV_TT‐RBD, has been optimized to incorporate a universal T helper cell epitope derived from tetanus‐toxin and is self‐adjuvanted with TLR7/8 ligands.Results: CuMV_TT‐RBD vaccine elicited a strong systemic RBD‐ and spike‐IgG and IgA antibodies of high avidity. Local immune response was assessed, and our results demonstrate a strong mucosal antibody and plasma cell production in lung tissue. Furthermore, the induced systemic antibodies could efficiently recognize and neutralize different variants of concern (VOCs).Conclusion: Our data demonstrate that intranasal administration of CuMV_TT‐RBD induces a protective systemic and local specific antibody response against SARS‐CoV‐2 and its VOCs.

Background: Several new variants of SARS‐CoV‐2 have emerged since fall 2020 which have multiple mutations in the receptor‐binding domain (RBD) of the spike protein. It is unclear which mutations affect receptor affinity versus immune recognition.Methods: We produced wild type RBD, RBD with single mutations (E484K, K417N, or N501Y) or with all three mutations combined and tested their binding to ACE2 by biolayer interferometry (BLI). The ability of convalescent sera to recognize RBDs and block their interaction with ACE2 was tested as well.Results: We demonstrated that single mutation N501Y increased binding affinity to ACE2 but did not strongly affect its recognition by convalescent sera. In contrast, single mutation E484K had almost no impact on the binding kinetics, but essentially abolished recognition of RBD by convalescent sera. Interestingly, combining mutations E484K, K417N, and N501Y resulted in a RBD with both features: enhanced receptor binding and abolished immune recognition.Conclusions: Our data demonstrate that single mutations either affect receptor affinity or immune recognition while triple mutant RBDs combine both features.

It is known that Müllerian ducts develop into the upper two-thirds of the vagina, uterus and Fallopian tubes ( Varner et al., 1985 x Varner et al., 1985 Varner, R.E., Younger, J.B., and Blackwell, R.E. Mullerian dysgenesis. J. Reprod. Med. 1985; 30: 443–450 PubMed | Google Scholar See all References Varner et al., 1985 ) and any alteration in the developmental sequence would lead to a wide spectrum of female reproductive tract abnormalities, called Müllerian duct abnormalities (MDA). The clinical presentation of MDA varies including congenital vagina and uterus defect (Mayer-Rokitansky–Küster–Hauser (MRKH) syndrome; Griffin et al., 1976 x Griffin et al., 1976 Griffin, J.E., Edwards, C., Madden, J.D., Harrod, M.J., and Wilson, J.D. Congenital absence of the vagina: The Mayer–Rokitansky–Kuster–Hauser syndrome. Ann. Intern. Med. 1976; 85: 224–236 Crossref | PubMed | Scopus (253) | Google Scholar See all References Griffin et al., 1976 ), uterine agenesis, unicornuate uterus, didelphic uterus, bicornuate uterus, septate uterus and vaginal septum ( Creatsas et al., 2010 x Creatsas et al., 2010 Creatsas, G., Deligeoroglou, E., and Christopoulos, P. Creation of a neovagina after Creatsas modification of Williams vaginoplasty for the treatment of 200 patients with Mayer–Rokitansky–Kuster-Hauser syndrome. Fertil. Steril. 2010; 94: 1848–1852 Abstract | Full Text | Full Text PDF | PubMed | Scopus (29) | Google Scholar See all References , Parelkar et al., 2009 x Parelkar et al., 2009 Parelkar, S.V., Gupta, R.K., Oak, S., Sanghvi, B., Kaltari, D., Patil, R.S., Prakash, A., and Shimoga, P. Laparoscopic management of persistent mullerian duct syndrome. J. Pediatr. Surg. 2009; 44: e1–e3 Abstract | Full Text | Full Text PDF | PubMed | Scopus (14) | Google Scholar See all References , Tiker et al., 2000 x Tiker et al., 2000 Tiker, F., Yildirim, S.V., Barutcu, O., and Bagis, T. Familial mullerian agenesis. Turk. J. Pediatr. 2000; 42: 322–324 PubMed | Google Scholar See all References ). It is often accompanied with other somatic malformations, such as renal agenesis (25%), skeletal anomalies (10–12%) and, more rarely, cardiac or middle ear anomalies ( Morcel et al., 2007 x Morcel et al., 2007 Morcel, K., Camborieux, L., and Guerrier, D. Mayer–Rokitansky–Küster–Hauser (MRKH) syndrome. Orp. J. Rare Dise. 2007; 2: 13 Crossref | PubMed | Scopus (186) | Google Scholar See all References Morcel et al., 2007 ). Patients with MDA are asymptomatic before preadolescence and would not be diagnosed with a uterine defect until the child-bearing period because of infertility, recurrent spontaneous abortion, premature birth, abnormal fetal presentation or cyclic pain ( Bernardini et al., 2009 x Bernardini et al., 2009 Bernardini, L., Gimelli, S., Gervasini, C., Carella, M., Baban, A., Frontino, G., Barbano, G., Divizia, T.M., Fedele, L., Novelli, A., Bena, F., Lalatta, F., Miozzo, M., and Dallapiccola, B. Recurrent microdeletion at 17q12 as a cause of Mayer-Rokitansky–Kuster-Hauser (MRKH) syndrome: two case reports. Orphanet J. Rare Dis. 2009; 4: 25 Crossref | PubMed | Scopus (71) | Google Scholar See all References , Biason et al., 2007 x Biason et al., 2007 Biason, L.A., De Filippo, G., Konrad, D., Scarano, G., Nazzaro, A., and Schoenle, E.J. WNT4 deficiency-a clinical phenotype distinct from the classic Mayer-Rokitansky–Kuster-Hauser syndrome: a case report. Hum. Reprod. 2007; 22: 224–229 Crossref | PubMed | Scopus (103) | Google Scholar See all References , Sadic et al., 2002 x Sadic et al., 2002 Sadic, S., Taskin, O., Sehirali, S., Mendilcioglu, I., önoglu, A.S., Kursun, S., and Wheeler, M.J. Complex mullerian malformation: report of a case with a hypoplartic non-carvitated uterus and two rudimentary horns. Hum. Reprod. 2002; 17: 1343–1344 Crossref | PubMed | Google Scholar See all References ). The incidence of MDA in infertile women is about 5% ( Nik-Zainal et al., 2011 x Nik-Zainal et al., 2011 Nik-Zainal, S., Strick, R., Storer, M., Huang, N., Rad, R., Willatt, L., Fitzgerald, T., Martin, V., Sandford, R., Carter, N.P., Janecke, A.R., Renner, S.P., Oppelt, P.G., Oppelt, P., Schulze, C., Brucker, S., Hurles, M., Beckmann, M.W., Strissel, P.L., and Shaw-Smith, C. High incidence of recurrent copy number variants in patients with isolated and syndromic Müllerian aplasia. J. Med. Genet. 2011; 48: 197–204 Crossref | PubMed | Scopus (56) | Google Scholar See all References Nik-Zainal et al., 2011 ).