Rational design of a SOCS1-edited tumor-infiltrating lymphocyte therapy using CRISPR/Cas9 screens.

Cell therapies such as tumor-infiltrating lymphocyte (TIL) therapy have shown promise in the treatment of patients with refractory solid tumors, with improvement in response rates and durability of responses nevertheless sought. To identify targets capable of enhancing the antitumor activity of T cell therapies, large-scale in vitro and in vivo clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 screens were performed, with the SOCS1 gene identified as a top T cell-enhancing target. In murine CD8+ T cell-therapy models, SOCS1 served as a critical checkpoint in restraining the accumulation of central memory T cells in lymphoid organs as well as intermediate (Texint) and effector (Texeff) exhausted T cell subsets derived from progenitor exhausted T cells (Texprog) in tumors. A comprehensive CRISPR tiling screen of the SOCS1-coding region identified sgRNAs targeting the SH2 domain of SOCS1 as the most potent, with an sgRNA with minimal off-target cut sites used to manufacture KSQ-001, an engineered TIL therapy with SOCS1 inactivated by CRISPR/Cas9. KSQ-001 possessed increased responsiveness to cytokine signals and enhanced in vivo antitumor function in mouse models. These data demonstrate the use of CRISPR/Cas9 screens in the rational design of T cell therapies.

Imidazo[1,5-a]quinoxalines as irreversible BTK inhibitors for the treatment of rheumatoid arthritis.

Imidazo[1,5-a]quinoxalines were synthesized that function as irreversible Bruton's tyrosine kinase (BTK) inhibitors. The syntheses and SAR of this series of compounds are presented as well as the X-ray crystal structure of the lead compound 36 in complex with a gate-keeper variant of ITK enzyme. The lead compound showed good in vivo efficacy in preclinical RA models.

Selective functional inhibition of JAK-3 is sufficient for efficacy in collagen-induced arthritis in mice.

OBJECTIVE: All gamma-chain cytokines signal through JAK-3 and JAK-1 acting in tandem. We undertook this study to determine whether the JAK-3 selective inhibitor WYE-151650 would be sufficient to disrupt cytokine signaling and to ameliorate autoimmune disease pathology without inhibiting other pathways mediated by JAK-1, JAK-2, and Tyk-2. METHODS: JAK-3 kinase selective compounds were characterized by kinase assay and JAK-3-dependent (interleukin-2 [IL-2]) and -independent (IL-6, granulocyte-macrophage colony-stimulating factor [GM-CSF]) cell-based assays measuring proliferation or STAT phosphorylation. In vivo, off-target signaling was measured by IL-22- and erythropoietin (EPO)-mediated models, while on-target signaling was measured by IL-2-mediated signaling. Efficacy of JAK-3 inhibitors was determined using delayed-type hypersensitivity (DTH) and collagen-induced arthritis (CIA) models in mice. RESULTS: In vitro, WYE-151650 potently suppressed IL-2-induced STAT-5 phosphorylation and cell proliferation, while exhibiting 10-29-fold less activity against JAK-3-independent IL-6- or GM-CSF-induced STAT phosphorylation. Ex vivo, WYE-151650 suppressed IL-2-induced STAT phosphorylation, but not IL-6-induced STAT phosphorylation, as measured in whole blood. In vivo, WYE-151650 inhibited JAK-3-mediated IL-2-induced interferon-gamma production and decreased the natural killer cell population in mice, while not affecting IL-22-induced serum amyloid A production or EPO-induced reticulocytosis. WYE-151650 was efficacious in mouse DTH and CIA models. CONCLUSION: In vitro, ex vivo, and in vivo assays demonstrate that WYE-151650 is efficacious in mouse CIA despite JAK-3 selectivity. These data question the need to broadly inhibit JAK-1-, JAK-2-, or Tyk-2-dependent cytokine pathways for efficacy.