Abstract 2696: Genetic and pharmacologic evaluation of the ubiquitin ligase CBL-B as a small-molecule, tumor immunotherapy target
Authors: Gosling Jennifa , Rountree Ryan , Taherbhoy Asad , Wang Chenbo , Cummins Thomas , Cohen Frederick , Tanaka Hiroko , Weiss Dahlia , Cardozo Mario , Karim Christopher , Tan May , Juan Joseph , Tenn-McClellan Austin , Soly Szerenke Kiss von , Sheung Julie , Boyle Kathleen , Dhamnaskar Ketki , Kurylo Katherine , Bruffey Jilliane , McKinnell Jennifer , Karr Dane , Christianson Andria , Vries Anne-Renee Van Der Vuurst de , Sivakumar Pallavur , Gallop Mark , Barsanti Paul A. , Saha Anjanabha , Bence Neil F. , Zapf Christoph W. 
Published: August 2019

E3 ubiquitin ligases play critical roles in directing cellular protein fate by controlling the specificity of ubiquitin conjugation to substrate proteins and targeting them for cellular relocalization or degradation by the ubiquitin proteasome system. The E3 ubiquitin ligase CBL-B is expressed in immune cell lineages and negatively regulates activity of the T-cell receptor (TCR) by imposing a requirement for a costimulatory signal to mount a productive immune response upon TCR engagement. Mice deficient in Cbl-b, and more specifically in the RING Zn-finger ligase domain of Cbl-b, demonstrate a tumor rejection phenotype mediated by CD8+ T cells (Paolino et al., JI, 2011). We have reproduced these results and demonstrate that Cbl-b deficient mice show enhanced anti-tumor activity. In addition, we show that CD4+ and CD8+ T cells from mice deficient in Cbl-b have 5 to 10-fold enhanced secretion of IL-2 and IFN γ when stimulated ex vivo. These data provide a genetic rationale for the development of a small molecule inhibitor of CBL-B ligase activity for use in patients with tumor-mediated immune suppression of effector T cells.

We have identified a series of small molecule inhibitors of CBL-B activity with biochemical potency at low nanomolar concentrations. CBL-B inhibitors increased cytokine secretion in vitro at low nanomolar concentrations, as measured by IL-2 and IFN γ secretion, in primary human and mouse T cells stimulated with CD3/CD28 or CD3 alone. The compounds also stimulated proliferation and elevated levels of the T cell surface activation markers CD25 and CD69. CBL-B inhibitors enhanced an antigen recall response in human PBMCs ex vivo, as measured by approximately 5-fold higher secretion of GM-CSF, TNF-α and RANTES, and demonstrated effects in an ex vivo model of exhausted T cell function.

Oral dosing of an optimized CBL-B inhibitor enhanced anti-CD3 stimulated T cell activation in mouse CD4+ and CD8+ T cells, demonstrating a dose proportional pharmacodynamic effect. Oral administration over 28 days in the syngeneic CT-26 tumor model was well tolerated and resulted in single agent tumor growth inhibition.

These data support the continued advancement of small molecule oral CBL-B inhibitors for future development in immuno-oncology.

Citation Format: Jennifa Gosling, Ryan Rountree, Asad Taherbhoy, Chenbo Wang, Thomas Cummins, Frederick Cohen, Hiroko Tanaka, Dahlia Weiss, Mario Cardozo, Christopher Karim, May Tan, Joseph Juan, Austin Tenn-McClellan, Szerenke Kiss von Soly, Julie Sheung, Kathleen Boyle, Ketki Dhamnaskar, Katherine Kurylo, Jilliane Bruffey, Jennifer McKinnell, Dane Karr, Andria Christianson, Anne-Renee Van Der Vuurst de Vries, Pallavur Sivakumar, Mark Gallop, Paul A. Barsanti, Anjanabha Saha, Neil F. Bence, Christoph W. Zapf. Genetic and pharmacologic evaluation of the ubiquitin ligase CBL-B as a small-molecule, tumor immunotherapy target [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2696.