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1208 DuoBody®-PD-L1x4–1BB (GEN1046) reverses T-cell exhaustion in vitro
  1. Jordan Blum1,
  2. Vanessa Spires1,
  3. Gregg Masters1,
  4. Christina Yu1,
  5. Saskia Burm2,
  6. Matt Hancock2,
  7. Omar Jabado1,
  8. Brandon Higgs1,
  9. Alexander Muik3,
  10. Kristina Nuermberger3,
  11. Friederike Gieseke3,
  12. Tahamtan Ahmadi1,
  13. Kate Sasser1,
  14. Ozlem Tureci3,
  15. Maria Jure-Kunkel1,
  16. Ugur Sahin3,
  17. Mark Fereshteh1 and
  18. Nora Pencheva2
  1. 1Genmab US, Princeton, NJ, USA
  2. 2Genmab BV, Utrecht, Netherlands
  3. 3BioNTech SE, Mainz, Germany


Background DuoBody®-PD-L1x4-1BB (GEN1046) is an investigational, potential first-in-class bispecific immunomodulatory antibody designed to elicit an anti-tumor immune response by simultaneous and complementary blockade of PD-L1 on tumor or immune cells and conditional 4-1BB stimulation on T cells and NK cells. Here we utilized a multi-omics approach to evaluate whether DuoBody-PD-L1x4-1BB could reverse T-cell exhaustion in vitro.

Methods An in vitro mixed lymphocyte reaction (MLR) assay was developed, optimized, and validated, where healthy donor T cells were exhausted by repeated stimulation with anti-CD3/CD28 beads prior to co-culturing with allogeneic LPS-matured dendritic cells. Publicly available single cell RNA sequencing (scRNAseq) datasets were harmonized across multiple solid-tumor indications (including treatment-naïve and anti-PD-1 and/or anti-CTLA-4 pretreated samples) and analyzed for co-expression of PD-1 and 4-1BB on various immune-cell subsets based on their transcriptome signatures.

Results In the T-cell exhaustion MLR assay, T cells showed increased expression of inhibitory receptors (e.g., TIM-3, LAG-3) and exhibited hyporesponsiveness for both proliferation and cytokine secretion upon restimulation with anti-CD3/CD28 beads, which was partially reversed by PD-1 blockade. DuoBody-PD-L1x4-1BB reinvigorated the exhausted T-cell response in vitro, as shown by restored IFNγ secretion. The effect of DuoBody-PD-L1x4-1BB in this assay was roughly two-fold higher to that of PD-1 blockade. When combined, DuoBody-PD-L1x4-1BB showed the potential to synergize with anti-PD-1 antibody treatment as cytokine secretion was further potentiated compared to each agent alone. More extensive molecular profiling from the T-cell exhaustion MLR assay will be presented. Using solid tumor public scRNAseq datasets, we demonstrated co-expression of 4-1BB and PD-1 on exhausted CD8+ T cells in the tumor microenvironment. Furthermore, in patients treated with agents that block the PD-1 pathway, an increase in exhausted CD8+ T cells expressing PD-1 was observed.

Conclusions PD-1 and 4-1BB are co-expressed on exhausted CD8+ T cells within the tumor microenvironment in solid tumors and T cell dysfunction may represent a potential resistance mechanism to checkpoint inhibitors (CPI). DuoBody-PD-L1x4-1BB restored IFNγ secretion by exhausted T cells in vitro more potently than PD-1 blockade, which could be further potentiated by the combination with an anti-PD-1 antibody. Together, these results support evaluation of DuoBody-PD-L1x4-1BB in the post-CPI setting and the combination of tumor-targeted 4-1BB co-stimulation with PD-1 checkpoint blockade for the treatment of solid tumors. DuoBody-PD-L1x4-1BB is currently being investigated in an ongoing phase 2 clinical trial in NSCLC patients who have progressed on prior CPI therapy (NCT05117242).

Acknowledgements These experiments were funded by Genmab A/S and BioNTech SE.

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