333 Targeting the apical intracellular checkpoint CISH unleashes T cell neoantigen reactivity and effector program

Abstract

BackgroundNeoantigen-specific T cells isolated from tumors have shown promise clinically but fail to consistently elicit durable tumor regression. Expression of the intracellular checkpoint CISH is elevated in human tumor infiltrating lymphocytes (TIL) and has been shown to inhibit neoantigen reactivity in murine TIL.MethodsTo explore CISH function in human T cells we developed a CRISPR/Cas9-based strategy to knockout (KO) CISH in human T cells with high-efficiency (>90%) and without detectable off-target editing.ResultsCISH KO in peripheral blood T cells enhanced proliferation, cytokine polyfunctionality, and cytotoxicity in vitro. To determine if CISH KO similarly enhances TIL function, we developed a clinical-scale, GMP-compliant manufacturing process for CISH disruption in primary human TIL. In process validation runs we achieved CISH KO efficiencies >90% without detectable off-target editing while maintaining high viability and expansion. Compared to WT controls, CISH KO in patient-derived TIL demonstrated increased proliferation, T cell receptor (TCR) avidity, neoantigen recognition, and unmasked reactivity to common p53 mutations. Hyperactivation in CISH KO TIL did not increase differentiation, suggesting that CISH KO may uncouple activation and differentiation pathways. Single cell profiling identifies a pattern of CISH expression inverse to key regulators of activation, and CISH KO in human TIL increases PD1 expression. Adoptive transfer of Cish KO T cells synergistically combines with PD1 inhibition resulting in durable tumor regression in mice, highlighting orthogonal dual cell surface and intracellular checkpoint inhibition as a novel combinatorial approach for T cell immunotherapy.ConclusionsThese pre-clinical data offer new insight into neoantigen recognition and serve as the basis for a recently initiated human clinical trial at the University of Minnesota (NCT04426669) evaluating inhibition of the novel intracellular immune checkpoint CISH in a CRISPR-engineered, neoantigen-specific T cell therapy for solid tumors. Updates from the clinical trial will be highlighted.Trial RegistrationNCT04426669

Epitranscriptomic Approach: To Improve the Efficacy of ICB Therapy by Co-Targeting Intracellular Checkpoint CISH

“…Cytokine-inducible SH2-domain containing protein (CISH or CIS) is one of the eighth members of SOCS family of proteins, recently gaining high attention due to its widespread regulatory role in cytokine signalling [170,171] and its involvement in more than 349-diseased (https://platform.opentargets.org/target/ENSG00000114737/associations; accessed on: 25 July 2021) [172][173][174] phenotypes. The therapeutic significance of 'CISH' can be evidenced by a recent clinical trial (NCT04426669, NCT03538613 by Intima Bioscience, UK; and ONKT102, ONKT103 and ONKT104 by ONK therapeutics, Ireland) targeting NK-cells, TILs and DCs for the treatment of broad range of metastatic cancers [1,2]. The so-called personalized medicine targeting 'CISH' in immune cells has shown promising effect in improving the efficacy of ICB-therapeutics [3,5].…”

“…Therefore, co-targeting CISH −/− TILs in combination with ICB-therapy would hold the potential to control tumor progression by improving the efficacy of ICB-antibodies as well as CD8 + T-cell effector function. A relevant human Phase-I/II clinical trial (NCT04426669, NCT03538613) targeting CISH −/− TILs in combination with ICB-therapy was proposed by Intima Bioscience, UK, until 2021𠄲2022 against wide range of tumor types and gastrointestinal cancer [ 1 , 2 , 115 ] ( Figure 8 B,E, Table 3 , Box-1).…”

“…Intima Bioscience is a UK-based Biotechnology Company, founded in 2021, conducting phase-I/II clinical trials (NCT04426669) against metastatic gastrointestinal (GIT) cancer patients by administering CISH-inactivated TILs by CRISPR/Cas9 system [ 1 ]. It is estimated to complete the trial by 31 October 2022 in collaboration with Masonic Cancer Center, University of Minnesota, USA [ 5 ].…”

“… NK-cells targeting intracellular checkpoint CISH: ONK therapeutics, Ireland, estimated to conduct Phase-I clinical trial (ONKT102, ONKT103 and ONKT104) by 2021𠄲2022 for the treatment of haematological malignancies (multiple myeloma and AML) and solid tumors (ovarian, NSCLC and breast cancers) [ 135 ]. TILs targeting intracellular checkpoint CISH: Intima Bioscience, UK, estimated to conduct Phase-I/II clinical trial (NCT04426669, NCT03538613) by 2021𠄲2022 for the treatment of wide range of tumor types and gastrointestinal cancer [ 1 , 2 , 5 ]. DCs targeting intracellular checkpoint SOCS-1: Military medical sciences, China, conducted Phase-I clinical trial (NCT01956630) in 2018 for the treatment of RAL [ 7 , 182 ].…”

“…Moreover, we have also emphasized co-targeting immune checkpoint proteins (PD-1 and PD-L1) along with intracellular checkpoint molecules (CISH, SOCS-1 and microRNAs) in enhancing the efficacy of ICB-therapeutics by combining immunotherapy. A recent human clinical trial NCT04426669, NCT03538613 [ 1 , 2 ] evidenced the success of targeting CISH/SOCS-1 in NK-cells [ 3 , 4 ], T-cells [ 5 , 6 ], and DCs [ 7 ] in further strengthening the efficacy of ICB-therapy against broad range of solid tumors and metastatic gastrointestinal cancers ( Table 1 , Table 2 and Table 3 ) and ( Figure 1 ).…”