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1354 CBL-B inhibition showed differentiated effects in a mixed lymphocyte reaction versus other immuno-oncology targeted approaches
  1. Yilin Qi,
  2. Jun Kuai,
  3. Yingzhi Bi,
  4. Huadong Sun,
  5. David Greco,
  6. Ken Carson,
  7. Timothy Reilly,
  8. Geraldine Harriman and
  9. Fang Wang
  1. Hotspot Therapeutics, Boston, MA, USA


Background The mixed lymphocyte reaction (MLR) mimics an immune reaction between T cells and antigen presenting cells and has been used to assess a variety of immune-oncology (I-O) agents as a potential predictor of clinical effects. Inhibition of the E3 ligase, Casitas B-Lineage Lymphoma Proto-Oncogene B (CBL-B), is a novel I-O approach shown to lower the threshold for antigen-specific T cell activation, even in the absence of co-stimulatory signaling or in the presence of an immunosuppressive environment. Moreover, genetic ablation of CBL-B and functional inactivation of its E3 ligase activity in mice or primary human T cells enhanced immune-mediated anti-tumor effects. Previously, Hotspot has disclosed a series of allosteric CBL-B inhibitors (CBL-Bi) with potent in vitro effects on T cells and NK cells and immune-mediated anti-tumor effects in vivo.

Methods A human allogenic MLR assay was applied to compare the effects of CBL-Bi and antibodies directed against PD1, CTLA4, LAG3, TIGIT and TIM3 on cytokine release and T cell proliferation, followed by evaluation of CBL-Bi effects on dendritic cells maturation and antigen specific CMV response.

Results CBL-Bi significantly enhanced interferon-gamma production comparable to anti-PD1 treatment, with an additional profound effect on CD8 T cell proliferation. Antibodies directed against CTLA4, LAG3, TIGIT and TIM3 had no effect on either endpoint in this assay format. The effects of CBL-Bi extended to CD4 T cells and interleukin-2 (IL-2) production; anti-PD1 also showed similar effects on IL-2. CBL-Bi effects were concentration-dependent, suggesting potential to optimize the extent of desired immune enhancement. The combination of CBL-Bi plus anti-PD1 demonstrated additive or possibly synergistic effects on both cytokine release and T cell proliferation. Combination of CBL-Bi plus other I-O agents did not show any enhancement. Mechanistically, CBL-Bi promoted immature dendritic cell activation and increased the sensitivity to antigen-specific T cell activation in a CMV challenge assay. An integrated literature review further suggested that the MLR assay has been generally correlated with clinical effects of I-O agents as monotherapy and/or in combination with anti-PD1.

Conclusions In summary, CBL-Bi had robust single agent effects on both cytokine release and T cell proliferation in the human MLR assay. These effects were differentiated from a range of other I-O mechanisms and CBL-Bi plus anti-PD1 showed substantial combination effects. These data support the potential for CBL-Bi to drive anti-tumor immunity in patients as both monotherapy and in combination with anti-PD1 standard of care therapy.

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