Functional Evaluation of Unique Anti-PDL1 Antibodies Generated through Single Plasma B Cell Cloning on the Beacon® Platform Versus a Standard Hybridoma Approach

PD-L1 is a key inhibitor of T cell activation that is often over-expressed in cancer to escape immune surveillance and promote tumor progression.  Blocking antibodies against PD-L1 or its receptor, PD-1, have shown significant clinical benefit in some patients with PD-L1 expressing tumors. Hence, there is great interest in generating therapeutic antibodies against these targets to counteract the immune suppression mechanism that tumors rely on for survival. Most of the anti-PD-L1 therapies in the clinic have been generated by standard hybridoma technology. We investigated whether superior anti-PD-L1 antibodies with greater diversity, affinity, and/or functional activity could be generated using single B cell cloning (BCC) which could circumvent a labor-intensive and time consuming process. Thus, we generated unique antibodies against PD-L1 via both methods. The Beacon® platform (Berkeley Lights, Inc., (BLI)) enabled screening of 33,377 antibody secreting plasma B cells (PCs) against PD-L1.  After penning CD138+ single PCs onto OptoSelectTM chips, we identified over 200 antibodies with binding to PD-L1, and of those, 35 antibodies blocked binding of PD-1 to PD-L1.  The hits were exported for antibody sequence recovery from single B cells and 24 hu-IgG4 chimeric antibodies were generated. For the traditional hybridoma approach, Balb/c and SJL mice were immunized with recombinant PD-L1 and 13,536 hybridoma cells were screened. We purified 44 hybridoma antibodies and 24 BCC antibodies which were characterized for FACS binding, receptor blocking, epitope binning & affinity measurements. Image based selection in the blocking assay enabled identification of 58% true blockers by Beacon with IC50s comparable to benchmarks. We found that 2 antibodies from Beacon and 3 antibodies from hybridoma had triple-digit pM affinity. Taken together, our data show that plasma B cells secreting functional antibody candidates can be rapidly identified on Beacon compared to several months for a hybridoma campaign, thus substantially accelerating the antibody discovery process.  Although the hybridoma approach allowed recovery of a greater number of blocking antibodies, B cell cloning on the Beacon platform enables greater access to the immune repertoire coupled with the ability to screen through robust functional assays up front, thus enabling identification of high affinity, potent anti-PD-L1 antibodies.  

Learning Objectives:

1. Functional assessment of clones on Beacon platform versus Hybridoma technology

2. Characterization of clones discovered by both methods

3. Comparison of clones by both methods against Benchmark antibodies