Correlates of protection for meningococcal surface protein vaccines: current approaches for the determination of breadth of coverage

Figures & data

Table 1. Current meningococcal surface protein vaccines and antigens

Vaccine	FHbp			NHBA		NadA		OMV
	Presentation	Subfamily and peptide	PubMLST peptide [variant]^a	Presentation	Peptide^a	Presentation	Peptide [variant]^a	Presentation	PorA subtype^a
4CMenB [25]	Non-lipidated single variant fused to 936 accessory protein	B24	ID1 [1]	Single variant fused to 953 accessory protein	2	Single variant	8 [2/3]	Produced from the New Zealand outbreak strain NZ 98/254	P1.7–2,4
MenB-FHbp [12]	Two lipidated variants	A05 & B01	ID45 [3] & ID55 [1]	NA		NA		NA

4CMenB = Bexsero®; FHbp = factor H binding protein; MenB-FHbp = Trumenba®, bivalent rLP2086; NA = not applicable; NadA = Neisserial adhesin A; NHBA = Neisserial heparin binding antigen; PorA = porin A. ^aPubMLST website located at https://pubmlst.org/

Figure 1. Phylogenetic tree for FHbp.

Phylogenetic tree for FHbp, adapted with permission from Ostergaard et al. [65]. FHbp peptides are grouped into subfamilies A and B [32]; or alternatively into three variant groups [12]. Colored circles indicate FHbp peptides included in 4CMenB [25] and MenB-FHbp [12] as well as that harbored by strains used in clinical trials for the serologic evaluation of 4CMenB [68,69] and MenB-FHbp [86]. The 4CMenB indicator strain used to evaluate FHbp-mediated immune responses expresses FHbp B24 [ID1] and is thus homologous to the vaccine antigen [25]. The four primary and 10 additional strains used to evaluate the immune response to MenB-FHbp express sequence-diverse FHbp peptides that are heterologous to the vaccine antigens [86]. The scale bar indicates phylogenetic distance using peptide sequence. FHbp = factor H binding protein; 4CMenB = Bexsero®; hSBA = serum bactericidal antibody using human complement; MenB-FHbp = Trumenba®, bivalent rLP2086.

Table 2. Breadth of coverage of meningococcal surface protein vaccines: factors impacting immunogenicity and the ability of induced antibody to provide protection across diverse strains

Parameter	Factors
Antigen presentation	Formulation and presentation of each antigen can impact the ability to induce cross-reactive antibodies to diverse peptide variants. For example, non-lipidated FHbp and lipidated FHbp have different immunological profiles.
Immune response	Individual antigenic response: Cross-reactivity of induced antibody to diverse peptides varies between individuals. Age-related response: Cross-reactivity of induced antibody to diverse peptides may be age-related with the lowest responses in the younger age groups.
Protein	Gene presence/absence: No coverage afforded to strains without the relevant gene or where incomplete/interrupted. Peptide diversity: No coverage afforded to strains expressing diverse protein peptides to which vaccine-induced antibody is not cross-reactive. Antibody cross-reactivity is influenced by antigen presentation and the individual and age-related immune response. Surface expression: No coverage afforded to strains without sufficient surface expression of the protein. Local epidemiology: Depending on the protein, variations in gene presence/absence, peptide diversity and surface expression may vary across geographies and over time effecting coverage.

FHbp = factor H binding protein.

Table 3. Meningococcal serogroup B strains frequently used in the serum bactericidal antibody assay for the analysis of 4CMenB

								MATS RP^b
	FHbp subfamily and peptide (ID)^a		NadA peptide [variant]		NHBA peptide		PorA VR2	FHbp (PBT = 0.021)	NadA (PBT = 0.009)	NHBA (PBT = 0.29)	Clonal complex
Indicator strains (corresponding antigen)^cd
NZ 98/254 (PorA)	B03 (ID14)		-		2		P1.2-4	0.044	<LLOQ	0.847	41/44
44/76-SL (FHbp)	B24 (ID1)		-		3		P1.16	1.006	<LLOQ	0.352	32
5/99 (NadA)	A26 (ID23)		3^e [2/3]		20		P1.2	<LLOQ	1.089	0.179	8
M10173 (NHBA)	A12 (ID24)		-		10		P1.16-3	N/R	N/R	N/R	41/44
Additional strains^c
M01 240101	B44 (ID15)		-		21		P1.15-11	N/R	N/R	N/R	269
M00 242922	B16 (ID4)		-		2		P1.4	N/R	N/R	N/R	41/44
M01 240364	A47 (ID31)		5^e [2/3]		29		P1.2	N/R	N/R	N/R	11
M01 240355	A47 (ID31)		12 [4/5]		18		P1.14	N/R	N/R	N/R	213

Bold type indicates RP>PBT.

FHbp = factor H binding protein; LLOQ = lower limit of quantification; MATS = Meningococcal Antigen Typing System; N/R = not reported; NadA = Neisserial adhesin A; NHBA = Neisserial heparin binding antigen; PBT = positive bactericidal threshold; PorA = porin A; RP = relative potency; VR = variable region; - = does not harbor a NadA gene.

^aPubMLST website located at https://pubmlst.org/

^b[82]

^c[24,25]

^d[69]

^eDifference between these two peptides is restricted to their leader peptides.

Table 4. Meningococcal serogroup B strains used throughout the clinical development of MenB-FHbp

Strain	FHbp subfamily and peptide (ID)^a	FHbp expression (MEASURE MFI)	Median MEASURE MFI of strains with same FHbp peptide (±1 SD)^b	Clonal complex
Primary strains
PMB80	A22 (ID19)	3127	2502 (1952; 3207)	41/44
PMB2001	A56 (ID187)	5002	5002^c	213
PMB2948	B24 (ID1)	6967	8457 (6599; 10,839)	32
PMB2707	B44 (ID15)	11,283	14,753 (11,511; 18,907)	269
Additional strains
PMB3010	A06 (ID47)	3370	3008 (2410; 3958)	461
PMB3040	A07 (ID21)	1379	1100 (858; 1409)	162
PMB824	A12 (ID24)	2540	2467 (1925; 3161)	35
PMB1672	A15 (ID25)	2995	2904 (2266; 3721)	103
PMB1989	A19 (ID16)	1934	1759 (1372; 2254)	8
PMB3175	A29 (ID30)	3839	5994 (4677; 7682)	32
PMB1256	B03 (ID14)	3976	2935 (2290; 3762)	41/44
PMB866	B09 (ID13)	2089	2275 (1775; 2916)	269
PMB431	B15 (ID252)	3785	4822 (3763; 6180)	41/44
PMB648	B16 (ID4)	2347	1966 (1557; 2558)	41/44

FHbp = factor H binding protein; MEASURE = Meningococcal Antigen Typing System; MFI = mean fluorescence intensity; SD = standard deviation.

^aPubMLST website located at https://pubmlst.org/

^bBased on 1263 MenB isolates, except for variant group A07, which was calculated from 1814 strains.

^cOnly one strain expressing A56; thus, no SD values calculated

Table adapted from [86]

Table 5. Human serum bactericidal antibody assay titers 1 month following two doses of 4CMenB on a 0–2-month schedule in adolescents 12–18 years of age against primary meningococcal serogroup B strains used throughout the MenB-FHbp clinical program [97,98]

Strain				% with composite hSBA titer (95% CI)^c
Strain ID	FHbp subfamily and peptide (ID)^a	Clonal complex	% with ≥4-fold rises in hSBA titer from pre- to post-vaccination (95% CI)^b	Pre-vaccination	Post-2nd dose
PMB80	A22 (ID19)	41/44	73.0 (60.3–83.4)	6.8 (1.8–16.4)	35.2 (22.7–49.4)
PMB2001	A56 (ID187)	213	37.5 (24.9–51.5)
PMB2948	B24 (ID1)	32	91.8 (81.9–97.3)
PMB2707	B44 (ID15)	269	37.3 (25.0–50.9)

CI = confidence interval; FHbp = factor H binding protein; hSBA = serum bactericidal antibody using human complement.

^aPubMLST website located at https://pubmlst.org/

^bn = 59–63

^cA composite hSBA titer is where a subject has a seroprotective hSBA titer against each of the four strains (≥1:8 for A56 (ID187), B24 (ID1), and B44 (ID15) and ≥1:16 for A22 (ID19)).

Table 6. Key factors influencing laboratory assays assessing the breadth of coverage of meningococcal surface protein vaccines

Factor	Outcome	Impacts results of
ELISA to determine specific IgG, IgM, or total concentrations	Protection from invasive meningococcal disease is mediated through functional SBA. ELISA measures ‘non-functional’ antibody which varies between subjects in addition to SBA and is therefore not suitable.	ELISA
hSBA methodology: Basic methodology and reagents used (e.g. agar overlay, dribble, colormetric assays), Assay validation and variation observed intra-assay, inter-assay, inter-operator, and inter-critical reagents, Complement source (endogenous or exogenous), Complement type (serum or plasma and any processing such as antibody removal or inclusion of additives such as magnesium, calcium, or sugars), Complement and critical reagent qualification (acceptance criteria applied and permitted variation between lots), Strain handling and assay growth conditions which affect strain susceptibility to lysis.	Differences in methodology directly influence hSBA results in different laboratories and impact the absolute hSBA titers determined. This mainly impacts the proportions achieving any titer cutoff applied (e.g. ≥1:4 or LLOQ) and GMTs. Methodologies generally do not influence measures of response such as ≥4-fold rises which self-reference from baseline hSBA titer except apart from when assay sensitivity and range are lost.	hSBA assay
Selection of strains for the hSBA assay considering: Low baseline positivity, Overall susceptibility to lysis, High protein expression.	Strain selection can impact hSBA assay results either increasing (high susceptibility and expression) or reducing (low baseline positivity, low susceptibility, and low expression) titers and the proportions achieving the titer cutoff applied (e.g. ≥1:4 or LLOQ) and GMTs. Strain selection generally does not influence measures of response such as ≥4-fold rises which self-reference from baseline hSBA titer except when assay sensitivity and range are lost.	hSBA assay
hSBA titer calculation: Timepoint used as reference for 50% killing threshold (e.g. T0 v T60), Reciprocal or interpolation of hSBA titer.	Use of T0 and interpolation result in increased hSBA titers.	hSBA assay
Antigenic presentation: Different presentation of the same protein vaccine antigen (e.g. lipidated FHbp v non-lipidated FHbp).	Presentation impacts immunogenicity and the cross-reactive profile of induced antibody to diverse protein peptides. Generalizations between different vaccine formulations must not be made.	hSBA assay and VE
Immune response: Differences among individuals’ responses, Age-related response, Antibody persistence.	Immunogenicity, cross-reactive profile of induced antibody, and antibody persistence may vary between individuals and age groups. Generalizations between individuals and age groups should not be made without evidence. The use of pooled samples in hSBA immunogenicity studies will not provide information on individual responses, which will vary and may also show increased killing due to synergy of different antibody populations.	hSBA assay and VE
Diversity of MenB strains/isolates: Proportions with and without uninterrupted protein antigen genes, Sequence diversity in proteins used as antigens, Surface expression of proteins.	Key drivers of breadth of coverage and must be collectively considered. e.g. hSBA or VE results against a strain expressing high amounts of a protein which is closely matched with the vaccine antigen may not provide information on breadth of coverage and may also overestimate overall immunogenicity and protection afforded.	hSBA assay and VE
Epidemiological differences in protein antigen gene presence, sequence diversity, and expression: Between countries or regions, Over time, Among different age groups.	Depending on the antigen and its immunogenicity these aspects may drive changes in the breadth of coverage and require consideration.	hSBA assay and VE
Application of the correlate of protection: Use of hSBA titer ≥1:4, Use of increase cutoffs (e.g. LLOQ).	An hSBA titer ≥1:4 is the accepted correlate of protection. An hSBA titer <1:4 does not necessarily indicate disease susceptibility, highlighting the conservative nature of the correlate. Use of cutoffs >1:4 is more conservative and results in a reduced sensitivity of determining vaccine responses.	hSBA assay
Evaluation of the immune response can be achieved by: Applying a titer cutoff (e.g. ≥1:4 or LLOQ), Fold rises (e.g. ≥4-fold), GMTs.	hSBA titers and proportions achieving a defined cutoff are influenced by many of the aspects covered in this table. Baseline seropositivity can interfere with the application of cutoffs. Baseline seropositivity has less impact on GMTs and measures of response which self-reference from the baseline hSBA titer except when assay sensitivity and range are lost.	hSBA assay
Overall vaccine immunogenicity may be driven by: Main antigens (e.g. FHbp, NadA, NHBA, PorA), Minor OMV antigens, Meningococcal accessory proteins, Synergy.	The contribution of antigenic components individually and collectively must be understood when inferring SBA results between strains. Failure to account will result in over- or underestimation of breadth of coverage.	hSBA assay
The representative nature of strain pools	Selection of strains to estimate breadth of coverage should represent diversity.	hSBA assay
MATS:	Provides a phenotypic binary ‘covered’ or ‘not covered’ outcome which does not account for differences in individuals’ responses or account for age-related 4CMenB immunogenicity. The conservative nature of PBT assignment, synergy from different antibody populations, and lower NadA in vitro expression may lead to underestimation of predicted coverage.	MATS
MEASURE:	Provides a binary phenotypic assessment on if sufficient FHbp is surface expressed to be susceptible to post-MenB-FHbp immune sera. Does not account for differences in individuals’ responses. The conservative nature of the threshold assignment may lead to lower predictions.	MEASURE
gMATS and MenDeVAR	4CMenB provides a binary genetic assessment of if an isolate is MATS ‘covered,’ ‘not covered,’ or unknown. For MenB-FHbp, MenDeVAR provides a binary genetic assessment if it would be MEASURE ‘positive’ or where hSBA data show killing of that FHbp peptide. These systems share the same characteristics of foundation MATS, MEASURE, or hSBA assays, e.g. do not account for differences in individuals’ responses. Final outcome is based on previous MATS or MEASURE database data and this will lead to different results for some strains than if the MATS or MEASURE assay was completed for that isolate. Scheme can only assess peptides where there is sufficient previous data in the database. MenDeVAR estimates for 4CMenB and MenB-FHbp cannot be compared as different schemes with different volumes of data are used.	gMATS and MenDeVAR

FHbp = factor H binding protein; gMATS = Genetic Meningococcal Antigen Typing System; GMT = geometric mean titer; hSBA = serum bactericidal antibody using human complement; IgG = immunoglobulin G; IgM = immunoglobulin M; LLOQ = lower limit of quantification; MATS = Meningococcal Antigen Typing System; MenDeVAR = Meningococcal Deduced Vaccine Antigen Reactivity; NadA = Neisserial adhesin A; NHBA = Neisserial heparin binding antigen; PorA = porin A; T₀ = time point zero (beginning of hSBA assay); T₆₀ = time point sixty (end of hSBA assay); VE = vaccine effectiveness.

Table

4CMenB	Bexsero®, MenB-4C
BAST	Bexsero Antigen Sequence Typing
cc	Clonal complex
CI	Confidence interval
ELISA	Enzyme-linked immunosorbent assay
FHbp	Factor H binding protein
gMATS	Genetic Meningococcal Antigen Typing System
GMT	Geometric mean titer
hSBA	Serum bactericidal antibody using human complement
IgG	Immunoglobulin G
IMD	Invasive meningococcal disease
LLOQ	Lower limit of quantification
MATS	Meningococcal Antigen Typing System
MEASURE	Meningococcal Antigen Surface Expression
MenACWY	Meningococcal serogroups A, C, W, and Y
MenB	Meningococcal serogroup B
MenB-FHbp	Trumenba®, bivalent rLP2086
MenC	Meningococcal serogroup C
MenDeVAR	Meningococcal Deduced Vaccine Antigen Reactivity
MenW	Meningococcal serogroup W
MenX	Meningococcal serogroup X
MenY	Meningococcal serogroup Y
NadA	Neisserial adhesin A
NHBA	Neisserial heparin binding antigen
OMV	Outer membrane vesicle
PBT	Positive bactericidal threshold
PorA	Porin A
RP	Relative potency
SBA	Serum bactericidal antibody
SD	Standard deviation
VR	Variable region

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