B cell fate following immunization : from memory B cells to plasma cells

Most approved successful human vaccines induce neutralizing antibody titers maintained above a given threshold for long-term protection against potential pathogen exposure. The pathogens targeted by these vaccines are antigenically stable and the relevant epitopes are immunogenic. In contrast, HIV-1 displays an enormous diversity in the circulating virus population and in each infected individual, in which the most relevant neutralizing epitopes are poorly exposed and thus less immunogenic HIV-1 is highly prone to immune escape, posing an extreme challenge for vaccine development. Elicitation of antibodies capable of neutralizing a broad range of HIV-1 strains and persist over time are likely to be required for an effective vaccine. This has focused the attention in the field on vaccine-elicited B cell responses against the HIV-1 envelope glycoproteins (Env), the only virally encoded target for neutralizing antibodies. Recent progresses in the design of soluble Env trimers that mimic the native HIV-1 spike have increased the interest in understanding vaccine-induced neutralizing antibody responses. In addition, the durability of vaccine-induced responses is poorly understood. Thus, a better understanding of how to modulate Env-induced responses by using different immunogens, immunization regimens and adjuvants is needed. In this thesis, I used rhesus macaques to investigate several of these questions.

In Paper I, we used an early generation Env trimer to evaluate whether the addition of a TLR-9 agonist to Matrix-M adjuvant would impact Env-specific immune responses. We demonstrated that the addition of the TLR-9 agonist had no measurable impact on the kinetics or durability of the B cell response, nor on the peripheral T cell response, the plasma neutralizing antibody activity or the control of viremia after challenge. In Paper II, we evaluated antibody responses elicited by new generation well-ordered HIV-1 trimers administered as soluble protein or conjugated to liposomes for multivalent display, both in the presence of Matrix-M adjuvant. We found that liposome-display resulted in superior germinal center (GC) responses and significantly improved neutralizing antibody activity compared to the soluble trimers. We then isolated monoclonal antibodies mediating autologous tier 2 virus neutralizing activity and demonstrated that these antibodies target the Env trimer apex using a lateral binding approach. In Paper III, we investigated plasma cell frequencies in the bone marrow (BM). Specifically, we evaluated whether longitudinal BM sampling would affect the frequency of total plasma cells in this compartment. We found this not to be the case; rather we observed intrinsic animal variation and that the frequency of plasma cells correlated with the age of the animals. In Paper IV, we described cell markers that better characterize BM plasma cells. We found that functional BM plasma cells that constitutively secrete IgG, IgA and IgM were double positive for CD138 and CD31. These markers allowed the distinction between bone marrow and peripheral plasma cells.

In conclusion, this thesis offers new information about several aspects of HIV-1 Env-induced B cell responses of direct relevance for vaccine development. This thesis also establishes methodology that can be used to further investigate vaccine-induced B cell responses, including in the BM compartment.

List of scientific papers

I. Martinez P, Sundling C, O'Dell S, Mascola J.R, Wyatt R.T, Karlsson Hedestam G.B. Primate immune responses to HIV-1 Env formulated in the saponin-based adjuvant AbISCO-100 in the presence or absence of TLR9 costimulation. Scientific Reports. 2015, 5 (8925): 1-11.
https://doi.org/10.1038/srep08925

II. Martinez-Murillo P, Tran K, Guenaga J, Lindgren G, Àdori M, Feng Y, Phad G.E, Vázquez Bernat N, Bale S, Ingale J, Dubrovskaya V, O’Dell S, Pramanik L, Spångberg M, Corcoran M, Loré K, Mascola J.R, Wyatt R.T and Karlsson Hedestam G.B. Particulate Array of Well-Ordered HIV Clade C Env Trimers Elicits Neutralizing Antibodies that Display a Unique V2 Cap Approach. [Accepted]
https://doi.org/10.1016/j.immuni.2017.04.021

III. Spångberg M, Martinez P, Fredlund H, Karlsson Hedestam G.B, Sundling C. A simple and safe technique for longitudinal bone marrow aspiration in cynomolgus and rhesus macaques. Journal of Immunological Methods. 2014, 408: 137-141.
https://doi.org/10.1016/j.jim.2014.05.004

IV. Martinez-Murillo P, Pramanik L, Sundling C, Hultenby K, Wretenberg P, Spångberg M, Karlsson Hedestam G.B. CD138 and CD31 double-positive cells comprise the functional antibody-secreting plasma cell compartment in primate bone marrow. Frontiers in Immunology. 2016, 7 (242): 1-10.
https://doi.org/10.3389/fimmu.2016.00242