Multiepitope Fusion Antigen: MEFA, an Epitope- and Structure-Based Vaccinology Platform for Multivalent Vaccine Development

A Polyvalent Adhesin–Toxoid Multiepitope-Fusion-Antigen-Induced Functional Antibodies against Five Enterotoxigenic Escherichia coli Adhesins (CS7, CS12, CS14, CS17, and CS21) but Not Enterotoxins (LT and STa)

“…Adhesin MEFA-IIb Presenting Epitopes of the Five ETEC Adhesins (CS7, CS12, CS14, CS17, and CS21), Two STa Toxoids, and an LT Epitope Was Constructed and Expressed Epitopes "AGSPVTRSDTTS", "GSNGQANNNDASQ", "TSGTAPSAGKYQ", "ADTQG-TAPEAGNY", and "VAMKDAYQRDGKYPDF" were identified in silico as the continuous immunodominant B-cell epitopes from the major subunits of adhesins CS7 (CsvA), CS12 (CswA), CS14 (CsuA), CS17 (CsbA), and CS21 (LngA), respectively. These five ETEC adhesin epitopes, two copies of full-length STa N12S toxoid (MNSSNYCCELCCSPACTGCY) and an epitope from LT A subunit (SPHPYEQEVSALTA) were presented on the backbone protein CfaB, the major subunit of CFA/I adhesin (Figure 1A,B), by replacing backbone epitopes, with the assistance of the MEFA vaccinology platform [16]. The resultant MEFA-IIb protein was confirmed in silico to be structurally stable (with an instability index of 39), with each adhesin and LT epitope antigenic (note: STa and STa toxoids are poorly immunogenic) and surface-exposed.…”

“…Therefore, eight continuous B-cell immunodominant epitopes from the CfaB backbone, which were predicted with the B-cell epitope prediction program BepiPred-2.0 (http://tools.iedb.org/bcell/help/#Bepipred-2.0 accessed on 26 September 2023), were substituted with two STa toxoids, an LT A epitope, and five adhesin epitopes. The resultant multiepitope fusion antigen was examined for epitope antigenicity, surface exposure, and epitope proposition with the IEBD program (www.iebd.org accessed on 26 September 2023), PyMol (www.pymol.org accessed on 26 September 2023), and Phyre2 (www.sbg.bio.ic.ac.uk/~phyre2 accessed on 26 September 2023), as we described previously [16,17]. The protein structure stability of the resultant adhesin MEFA was estimated with ExPASy (www.expasy.org accessed on 26 September 2023).…”

“…By applying an epitope-and structure-based multiepitope fusion antigen (MEFA) vaccinology platform [16], we initially constructed a polyvalent protein immunogen to induce cross-protective immunity against these five ETEC adhesins (CS7, CS12, CS14, CS17, and CS21) and found that this new MEFA protein antigen (termed as adhesin MEFA-II, to be differentiated with CFA/I/II/IV MEFA) is broadly immunogenic and induces functional antibodies against the five targeted ETEC adhesins as well as ETEC STa toxin [17]. Adhesin MEFA-II used adhesin CFA/I major subunit CfaB as the backbone to present epitopes from the major subunits of these five adhesins (CsvA of CS7, CswA of CS12, CsuA of CS14, CsbA of CS17, and LngA of CS21) as well as an STa toxoid (STa N12S ), as a polyvalent protein immunogen.…”

Intramuscularly Administered Enterotoxigenic Escherichia coli (ETEC) Vaccine Candidate MecVax Prevented H10407 Intestinal Colonization in an Adult Rabbit Colonization Model

“…Toxoid fusion protein 3xSTa N12S -mnLT R192G/L211A , which has three copies of STa toxoid STa N12S genetically fused to double mutant LT toxoid monomer mnLT R192G/L211A (a double mutant LT A subunit fused to an LT B subunit as a single polypeptide) ( 20 ), induces antitoxin antibodies neutralizing enterotoxicity of both ETEC toxins and, more importantly, protects against STa- or LT-mediated clinical diarrhea in a pig challenge model ( 17 , 20 – 22 ). ETEC adhesin CFA/I/II/IV MEFA protein, an epitope- and structure-based multiepitope fusion antigen protein ( 23 ), presents functional epitopes of seven ETEC adhesins, CFA/I, CS1, CS2, CS3, CS4, CS5, and CS6, on CFA/I adhesin major subunit CfaB backbone ( 24 ). This CFA/I/II/IV MEFA protein induces broad anti-adhesin antibodies that inhibit adherence of E. coli or ETEC strains expressing any of the seven target adhesins (CFA/I, CS1 to CS6) and significantly reduces ETEC strain B7A (CS6, STa, LT) colonization at rabbit small intestines ( 24 – 26 ).…”

A polyvalent multiepitope protein cross-protects against Vibrio cholerae infection in rabbit colonization and passive protection models

“…In this study, we constructed a polyvalent cholera protein immunogen by applying an epitope- and a structure-based vaccinology platform called multiepitope fusion antigen (MEFA) ( 33 ), then characterized the broad immunogenicity of the polyvalent MEFA protein, and examined protection from the MEFA protein–induced antibodies against V. cholerae intestinal colonization and clinical diarrhea preclinically. Mice immunized with this polyvalent protein were examined for antibody responses to the virulence factors targeted by the immunogen, and the derived antibodies were then evaluated for in vitro functions, including prevention of adherence from different V. cholerae serogroup strains, neutralization of CT enterotoxicity, blocking of CT binding to host receptor GM 1 , inhibition of vibrio motility, vibriocidal activity, and the activity against hemolysis.…”