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698 PD-L1 targeted CD28 costimulatory bispecific antibodies enhance T cell activation in solid tumors
  1. Veronica Zeng1,
  2. Gregory Moore2,
  3. Juan Diaz1,
  4. Christine Bonzon1,
  5. Kendra Avery1,
  6. Ruschelle Love1,
  7. Matthew Dragovich1,
  8. Rumana Rashid1,
  9. Irene Leung1,
  10. Michael Hackett1,
  11. Jing Qi1,
  12. Charles Bakhit1,
  13. Umesh Muchhal1,
  14. Norman Barlow1,
  15. John Desjarlais1 and
  16. Michael Hedvat1
  1. 1Xencor, Inc., Monrovia, CA, United States
  2. 2Xencor, Inc, Monrovia, CA, United States


Background T cells in the tumor microenvironment require T cell receptor (TCR) /major histocompatibility complex engagement and costimulatory receptor engagement to achieve complete activation. Tumor cells lack expression of CD28 ligands, so we hypothesized that activation of CD28 signaling at the T cell /tumor cell interface could enhance anti-tumor activity. We designed PD-L1 x CD28 bispecific antibodies that conditionally costimulate CD28 only in the presence of PD-L1 and TCR engagement. As PD-1/PD-L1 signaling has been shown to directly inhibit CD28 costimulation, this novel bispecific antibody can promote CD28 costimulation while simultaneously preventing the suppression of the same signal.

Methods We designed a set of stability-optimized anti-CD28 antibodies that can be paired with anti-PD-L1 antibodies to engage both PD-L1 and CD28 monovalently using Xencor's XmAb® 1+1 bispecific antibody platform. In vitro T cell activation with these bispecifics was measured by T cell proliferation, cytokine production, and cytotoxicity, in co-cultures of human cancer cell lines mixed with primary human CD3-stimulated T cells. In vitro activity was validated in a Cytomegalovirus (CMV) recall assay measuring CMV+ T cell proliferation of CMV+ peripheral blood mononuclear cells (PBMC) co-cultured with cancer cell lines ectopically treated with CMV-pp65-derived peptide. In vivo activity was determined with hCD28 humanized mice inoculated with MC38 tumors stably expressing hPD-L1-antigen. Finally, safety, tolerability, and pharmacodynamics of PD-L1 x CD28 were determined in cynomolgus monkeys.

Results PD-L1 x CD28 bispecifics were generated by incorporating an anti-PD-L1 mAb capable of blocking PD-1/PD-L1 interaction and anti-CD28 single-chain fragment variable covering a range of affinities. PD-L1 x CD28 antibodies enhanced T cell degranulation, cytokine secretion, and cancer cell cytotoxicity in concert with CD3 stimulation only in the presence of PD-L1. PD-L1 x CD28 enhanced proliferation of CMV+ T cells recognizing cancer cells loaded CMV-pp65-derived peptide. In hCD28 mice inoculated with MC38 tumors expressing hPD-L1, PD-L1 x CD28 inhibited tumor growth significantly greater than an anti-PD-L1 antibody alone. PD-L1 x CD28 was well tolerated in cynomolgus monkeys.

Conclusions PDL1 x CD28 bispecific antibodies show promising anti-tumor activity and warrant further development.

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