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844 A trispecific ROR1 x CD3 T cell engager mediates in vitro tumor cell killing and in vivo tumor eradication
  1. Bithi Chatterjee,
  2. Daniel Snell,
  3. Daniel Snell,
  4. Christian Hess,
  5. Matthias Brock,
  6. Fabio Spiga,
  7. Alexandre Simonin,
  8. Tea Gunde,
  9. Stefan Warmuth,
  10. Christopher Weinert,
  11. Nicole Bassler,
  12. Niels Kirk,
  13. Nina Schumacher,
  14. Dana Mahler,
  15. Yasemin Yaman,
  16. Bettina Bommer,
  17. Giorgio Gambino,
  18. Noreen Giezendanner,
  19. Benjamin Kuettner,
  20. Naomi Flueckiger,
  21. Robin Heiz,
  22. Sandro Wagen,
  23. Dania Diem,
  24. Julia Zeberer and
  25. David Urech
  1. Numab Therapeutics, Wadenswil, Switzerland


Background Receptor tyrosine kinase-like orphan receptor 1 (ROR1) is expressed on a variety of difficult to treat solid and hematological malignancies. Several therapeutic concepts targeting ROR1 are currently in clinical studies, including antibody-drug conjugates (ADCs), chimeric antigen receptor engineered T cells, as well as a bispecific T cell engager. In contrast to ADCs, T cell engagers have the capacity to induce tumor cell depletion irrespective of tumor cell mitotic activity. For the therapy of ROR1 expressing tumors, we engineered a T cell engager with prolonged half-life to support convenient administration schemes.

Methods NM32-2668, a ROR1-targeting T cell engager with prolonged serum half-life was engineered by joining three humanized rabbit antibody variable region (Fv) fragments specific for ROR1, CD3ɛ, and serum albumin, into our tri-specific scMATCHTM3 format. Each Fv fragment was stabilized using the ʎ-capTM technology. NM32-2668 was tested in assays for specific tumor lysis, induction of T cell proliferation, and cytokine release. These studies were performed using human T cells co-cultured with tumor cell lines and human tumor samples expressing various levels of ROR1. In vivo xenograft mouse studies were conducted using a human mantle cell lymphoma model in NCG mice engrafted with human PBMCs.

Results Here we report the design and the promising preclinical activity of the scMATCHTM3 ROR1/CD3/hSA T cell engager NM32-2668 in vitro and in vivo. Importantly, we demonstrate potent and specific cytotoxic activity in the sub-nanomolar range on tumor cell lines expressing different levels of ROR1. NM32-2668 also mediates ROR1 dependent T cell activation and cytokine release. We observe robust tumor cell killing activity of NM32-2668 over an extended time period and at multiple ratios of effectors to targets in a real time imaging-based cytotoxicity assay. This molecule also mediates T cell proliferation in response to target cell binding. NM32-2668 mediates in vitro lysis of CLL patient tumor cells, T cell activation, and cytokine release, with minimal IL-6 involvement. In an in vivo mantle cell lymphoma model (Jeko-1) engrafted with human PBMCs, we observe tumor regression and eradication.

Conclusions Collectively, these data demonstrate robust anti-tumor efficacy by NM32-2668, a scMATCHTM3 ROR1/CD3/hSA. Our results demonstrate that NM32-2668 promotes ROR1 dependent T cell activation and proliferation, as well as T cell-mediated tumor cell lysis. The activity of NM32-2668 has the potential to provide significant benefit to patients with ROR1+ malignancies on a convenient dosing schedule. We intend to rapidly progress NM32-2668 to clinical development.

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