Primary Antigen-Specific B Cells: A Novel Approach to Cellular-Based Immunotherapy.

Background: The potential for CD40 ligand (CD40L)-activated B cells to serve as antigen-presenting cells (APC) for cell-based immunotherapy has been suggested. Unlike dendritic cells (DC), CD40L-activated B cell populations are readily expandable in vitro. In addition, antigen-specific B cells may efficiently uptake, process, and present cognate protein antigens. Nevertheless, important questions regarding the relative efficacy of CD40L-activated B cells as cell-based vaccines remain. Here, we exploited the unique ability of B cells to uptake antigen through their B cell receptor (BCR) and the propensity for CD40L-activated B cells, including antigen-specific clones, to grow in culture and to process cognate protein antigens to determine if CD40L-activated B cells represent a suitable substitute for dendritic cells for cell-based immunotherapy.

Methods: As a head to head comparison between CD40L-activated B cells and mature DC, CD40L-activated B cells and bone marrow-derived DC were pulsed with MHC II- or MHC I-restricted self protein-derived (MOG; MBP) peptides and tested for their ability to induce proliferation of CD4⁺ or CD8⁺ clones. To compare processing and presentation of foreign protein antigens, C57BL/6 mice were immunized with 200 mg NP-BSA or an equivalent volume of PBS emulsified in CFA, sacrificed 10 days later and splenocytes obtained to generate antigen-specific CD40L-activated B cells and T cells. Bone marrow cells from PBS/CFA immunized mice were used to generate DCs. CD40L-activated (antigen-specific) B cells and DC were pulsed with NP-BSA, NP-CGG, or BSA and assayed for their ability to induce proliferation of primary T cells.

Results: B cell populations were readily expanded by culture on CD40L transfected L cells. CD40L stimulation significantly up-regulated MHC class I and II expression and induced expression of CD80 and CD86 to levels similar to those detected on mature DCs. CD40L-activated B cells were comparable to DCs when presenting MHC class I- or II-restricted self-peptides to T cell clones. When presenting cognate protein antigen (NP-BSA or BSA) to primary T cells, CD40L-activated B cells from NP-BSA immunized mice were as efficient as DC, both of which induced a 13–15 fold increase in T cell proliferation. To determine if the hapten moiety is sufficient to increase antigen up-take and presentation, DCs and CD40L-activated B cells from NP-BSA immunized mice were pulsed with NP-CGG and used as APC for T cells from NP-BSA immunized mice. DCs induced significant responses comparable to those seen with BSA and NP-BSA. Activated B cells from NP-BSA-immunized mice induced significantly higher responses to NP-CGG than activated B cells from control PBS/CFA “immunized” mice, although these responses were lower than those generated with dendritic cells.

Conclusion:

1. CD40L-activated B cells can be readily expanded in vitro

2. and significantly up-regulate co-stimulatory molecules CD80 and CD86 to levels comparable to mature DCs,

3. CD40L-activated B cells present MHC class I- and II-restricted self-peptides to T cell clones as efficiently as mature DCs,

4. Antigen-primed B cells are as efficient at presenting cognate protein antigens as DCs, Immunization with a hapten-carrier is sufficient to induce hapten-specific B cells which, when activated with CD40L, effectively present unrelated neoantigens conjugated with the hapten.

The data suggest that CD40L-activated B cells represent an important alternative APC for immunotherapy, particularly when previously educated to protein or haptenic determinants.