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751 Neo-X-Prime bispecific antibodies targeting CD40 and tumor antigens promote cross-presentation of tumor exosome-derived neoantigen and induce superior anti-tumor responses compared to CD40 mAb
  1. Karin Hagerbrand1,
  2. Mattias Levin1,
  3. Laura Von Schantz1,
  4. Laura Varas1,
  5. Anna Säll1,
  6. Adnan Deronic1,
  7. Anette Sundstedt1,
  8. Lill Ljung1,
  9. Karin Barchan1,
  10. Doreen Werchau1,
  11. Anna Rosén1,
  12. Barnabas Nyesiga1,
  13. Eva Lindqvist1,
  14. Mia Thagesson1 and
  15. Peter Ellmark2
  1. 1Alligator Bioscience, Lund, Sweden
  2. 2Alligator Bioscience AB, Lund, Sweden


Background Alligator's Neo-X-Prime platform aims to enable antigen presenting cells to efficiently enhance priming of tumor neoantigen-specific T cells with the goal of overcoming PD-1 resistance in certain tumor types. We hypothesize that binding of a CD40 x TAA bispecific antibody (bsAb) to CD40 on dendritic cells (DCs) and a tumor-associated antigen (TAA) on tumor exosomes or tumor debris leads to (i) activation of the DC, (ii) uptake of the tumor material, (iii) cross-presentation of tumor-derived neoantigen (present in exosomes or debris) and, iv) priming of tumor neoantigen-specific T cells, resulting in an increased quantity and/or quality of the tumor-targeting T cell pool.

Methods Functionality was evaluated in vitro using CD40 reporter cells and monocyte-derived DCs, co-cultured with cells expressing TAA. Further, co-localization of TAA-expressing cellular debris with a CD40-expressing human B cell line in the presence of bsAbs was assessed using live cell imaging. In vivo, anti-tumor efficacy and immunological memory were assessed in human CD40 transgenic (hCD40tg) mice bearing MB49 bladder carcinoma tumors transfected with human TAA or controls. T cells isolated from OVA-specific TCR-transgenic mice were used to evaluate the effect of Neo-X-Prime bsAbs on antigen-specific T cell expansion in the presence of hCD40tg DCs and exosomes from MB49 tumors transfected with both human TAA and OVA using flow cytometry.

Results Using CEA as a highly expressed TAA, we have developed lead Neo-X-Prime CD40-CEA bsAbs engineered to achieve an optimal profile. Further, using Neo-X-Prime concept molecules targeting EpCAM, we have demonstrated the ability to mediate co-localization of tumor debris and CD40 expressing antigen presenting cells that is dependent on the receptor density of the TAA. We have further shown that addition of Neo-X-Prime bsAbs to a co-culture of murine DCs, T cells and tumor-derived exosomes induces increased expansion of model neoantigen-specific T cells. In vivo, Neo-X-Prime bsAbs display a potent, TAA-dependent anti-tumor effect that is superior to CD40 mAbs. Cured mice develop a broad immunological memory that is not dependent on expression of the TAA. The tumor-localizing property of Neo-X-Prime bsAbs also shows potential for improved safety compared to CD40 monospecific antibodies.

Conclusions Neo-X-Prime bsAbs have the potential to tumor-selectively target CD40-expressing antigen-presenting cells to mediate an expansion of the tumor-specific T cell repertoire, resulting in increased T cell infiltration and potent anti-tumor effects.

Ethics Approval All experiments were performed after approval from the Malmö/Lund Animal Ethics Committee.

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