Research ArticleINFECTIOUS DISEASES

Clonal selection drives protective memory B cell responses in controlled human malaria infection

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Science Immunology  16 Feb 2018:
Vol. 3, Issue 20, eaap8029
DOI: 10.1126/sciimmunol.aap8029
  • Fig. 1 Molecular and functional characterization of antibodies from PfCSP-reactive memory B cells and plasmablasts.

    (A) PfSPZ-CVac immunization scheme (9). Blood samples were taken at day 7 (d7) after Pf exposure I, II, and III. (B to F) Ig gene sequence analysis of PfCSP-reactive memory B cells and plasmablasts from all (B to D) or individual (E and F) donors, as indicated. (B) Mean and SEM IGH isotype frequency. (C) IGHV absolute SHM counts and mean (red lines). (D) Mean and SEM ratio of replacement (R) to silent (S) SHM in CDRs compared with FWRs. (E) Clonal expansion. (F) Clonal expansion and diversity (day 7 after exposure III). Individual cell clusters are colored; unique sequences are white (top). Corresponding subsampling analysis (bottom). Dots represent independent analyses. Red lines show mean. Blue lines indicate observed values as in (E). (G to J) Recombinant monoclonal PfCSP memory B cell antibodies from eight donors at day 7 after Pf exposure I, II, and III. Representative optical density (OD) (G) and AUC values from PfCSP (H) and NANP (I) ELISA. Red and blue lines in (G) indicate positive (2A10) and negative (mGO53) controls. Antibody sporozoite inhibitory activity in traversal assays (J). (K to M) Affinity, frequencies (K), and kinetic on-rate (L) and off-rate (M) constants of NANP-reactive antibodies at day 7 after inoculation II and III. (H to J, L, and M) Red lines indicate mean. Data are representative of at least two independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, two tailed Mann-Whitney test (H to J, L, and M) and χ2 test (K). n numbers are provided in tables S1 and S2. ns, not significant.

  • Fig. 2 Inefficient affinity maturation.

    (A to C) NANP5 affinity for clonally related antibodies (closed circles) from the indicated clusters that do not show affinity maturation (A) including the predicted germline precursor (open circles) (B) and for B cell cluster antibodies, where SHM affects their NANP5 affinity (C). Data are representative of at least two independent experiments. ND, not detectable.

  • Fig. 3 Computational model of the B cell response to complex antigens.

    (A and B) Schemes of GC (A) and key mutation models (B). (C) Mean effect of single key mutations on log Kd for different nkey values. (D) Clonal composition of one exemplary GC (left) and mean cell and clone numbers of 500 GCs (right) over time. Colors indicate different clones. Inset: Affinity of individual memory cells stemming from the GC-dominating red clone over mutational state. (E) Left: Affinity of memory B cells produced by the GC reaction in (D). Colors indicate clones. Right: Affinity of GC memory output over time for different nkey values. Mean of 500 simulations, shaded area = 1 SD. (F) Dynamics of memory cell affinity and clonal diversity (normalized Shannon entropy) in a simulation of the experimental trial protocol (Fig. 1A). Mean of 50 simulations, shaded area = 1 SD. (G) Clonal composition of GCs after each infection. Clonally expanded high-affinity clones (red) seed and expand in successively more GCs after the second and third infection. (H) Sensitivity of the mean affinity (Kd) of the memory B cell pool after three infections to changes in nkey, GC peak size, GC lifetime, and antigen dose. Means of 50 simulations, error bars = 1 SD. Default values are highlighted. (I to L) Affinity distribution in GC precursor cells at day 0 (left) and corresponding memory B cells after three infections at day 126 (right). (I) Default conditions for nkey = 10 with a diverse precursor repertoire and natural mutation rate of 0.003 per codon and division. (J) As in (I), but with mutation rate of zero (no SHM possible). (K) As in (I), but limited repertoire without high-affinity GC precursors. (L) As in (I), but for a lower complexity antigen of nkey = 3. Memory B cells with equal, higher, or lower affinity than their corresponding GC precursor are shown in gray, green, and red, respectively. Ab, antibody.

  • Fig. 4 Recruitment of potent germline antibodies after repeated Pf exposure.

    (A) Antibody affinity versus SHM as predicted by the mathematical model (Fig. 3). Red and gray fillings indicate antibodies from memory B cells derived from clones that entered the Pf response as naïve or preexisting memory B cells, respectively. (B to G) Ig gene features of memory B cell antibodies from eight donors. PfCSP ELISA AUC (B) and Pf sporozoite inhibition activity (C) versus total IGHV plus IGKV/IGLV SHM counts. Dotted lines indicate germline antibodies. 2A10 (red) and mGO53 (blue) are shown as positive and negative controls, respectively. (D and E) PfCSP ELISA reactivity (AUC) versus IGHV family usage (D) and Igκ CDR3 (KCDR3) amino acid (aa) length (E). (F) NANP affinity for antibodies with 8– and 9–amino acid–long KCDR3s. (G) Frequencies of IGHV-IGKV gene association for antibodies with high (<10−7 M) and low (>10−7M) NANP affinity and 8– or 9–amino acid–long KCDR3. n indicates number of tested antibodies. Donors with observed antibody gene associations are listed. (H) Parasite-free mice after passive immunization and Pb-PfCSP infection. n = 5 per group. (I) Quality of PfCSP memory B cell antibodies from the indicated donors as measured by ELISA (AUC). n numbers are as shown in table S1. (J) Principal components analysis based on mean degree of clonal expansion after three Pf exposures (Fig. 1F, subsampling), frequency of antibodies expressing IGHV3 (fig. S6B), and frequency of antibodies with 8–amino acid–long KCDR3 (fig. S6C). (D to F and I) Red lines indicate mean. Data are representative of at least two independent experiments. ****P < 0.0001, two-tailed Mann-Whitney test.

Supplementary Materials

  • immunology.sciencemag.org/cgi/content/full/3/20/eaap8029/DC1

    Fig. S1. Anti-PfCSP response.

    Fig. S2. PfCSP memory B cell Ig gene sequence analysis and antibody function.

    Fig. S3. Inefficient affinity maturation over repeated Pf exposure.

    Fig. S4. Influence of GC seeder cell frequencies and antigen complexity (nkey) on clonal evolution within individual GCs.

    Fig. S5. Avalanche effect over three successive infections in an exemplary system of 10 different GC sites.

    Fig. S6. Repertoire and Ig gene feature analysis of antibodies from PfCSP memory B cells and plasmablasts.

    Table S1. Number of sequenced and cloned PfCSP-reactive memory B cell antibodies.

    Table S2. Number of sequenced plasmablast antibodies.

    Table S3. Clonally expanded PfCSP-reactive memory B cell clusters.

    Table S4. Simulation parameters, binding model, GC simulation dynamics.

    Table S5. PfCSP-reactive memory B cell antibodies with 8–amino acid–long KCDR3.

  • Supplementary Materials

    Supplementary Material for:

    Clonal selection drives protective memory B cell responses in controlled human malaria infection

    Rajagopal Murugan, Lisa Buchauer, Gianna Triller, Cornelia Kreschel, Giulia Costa, Gemma Pidelaserra Martí, Katharina Imkeller, Christian E. Busse, Sumana Chakravarty, B. Kim Lee Sim, Stephen L. Hoffman, Elena A. Levashina, Peter G. Kremsner, Benjamin Mordm?ller, Thomas Höfer,* Hedda Wardemann*

    *Corresponding author. Email: t.hoefer{at}dkfz-heidelberg.de (T.H.); h.wardemann{at}dkfz.de (H.W.)

    Published 16 February 2018, Sci. Immunol. 3, eaap8029 (2017)
    DOI: 10.1126/sciimmunol.aap8029

    This PDF file includes:

    • Fig. S1. Anti-PfCSP response.
    • Fig. S2. PfCSP memory B cell Ig gene sequence analysis and antibody function.
    • Fig. S3. Inefficient affinity maturation over repeated Pf exposure.
    • Fig. S4. Influence of GC seeder cell frequencies and antigen complexity (nkey) on clonal evolution within individual GCs.
    • Fig. S5. Avalanche effect over three successive infections in an exemplary system of 10 different GC sites.
    • Fig. S6. Repertoire and Ig gene feature analysis of antibodies from PfCSP memory B cells and plasmablasts.
    • Table S1. Number of sequenced and cloned PfCSP-reactive memory B cell antibodies.
    • Table S2. Number of sequenced plasmablast antibodies.
    • Table S3. Clonally expanded PfCSP-reactive memory B cell clusters.
    • Table S4. Simulation parameters, binding model, GC simulation dynamics.
    • Table S5. PfCSP-reactive memory B cell antibodies with 8–amino acid–long KCDR3.

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    Other Supplementary Material for this manuscript includes the following:

    Files in this Data Supplement:

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