P392: PHARMACOLOGICAL INHIBITION OF SYK CONFERS ANTI-PROLIFERATIVE AND NOVEL ANTI-TUMOR IMMUNE RESPONSES IN AML

Abstract

Background: Spleen tyrosine kinase (SYK) acts as a key integrator of signals from cell surface receptors containing an immunoreceptor tyrosine-based activation motif to boost cellular proliferation. In acute myeloid leukemia (AML), SYK serves as a relay to an oncogenic transcriptional regulatory network linked to NPM1, HOXA9 and MEIS1. The selective, orally bioavailable SYK inhibitor entospletinib (ENTO) has demonstrated clinical activity and tolerability in HOXA9/MEIS1-driven AML. ENTO is currently being investigated in a global Phase 3 trial, AGILITY (NCT05020665), in combination with intensive induction/consolidation chemotherapy in patients (pts) with treatment-naive NPM1-mutated (NPM1m) AML. Lanraplenib (LANRA) is a next-generation SYK inhibitor with similar potency and selectivity to ENTO but with more favorable pharmacologic properties that is currently being evaluated in combination with gilteritinib in pts with relapsed or refractory FLT3-mutated AML (NCT05028751). Aims: To investigate the effects of ENTO and LANRA in T-cell responses in AML. Methods: All pt-derived bone marrow (BM) and peripheral blood samples were obtained in accordance with IRB (OHSU#004422) approval. Results from gene expression profiling studies (RNA sequence [RNA-seq]) in bone marrow/peripheral blood mononuclear cells (MNCs) derived from 152 pts with AML, for which matching NPM1 status and ex vivo sensitivity to ENTO was available, previously published by Tyner et al (2018), were reanalyzed using trimmed mean of M values normalization and differential expression using edgeR. ENTO/LANRA sensitivity was assessed ex vivo after 72 hours in culture at 37°C by MTS assay. T-cell functional assays were performed by culturing MNCs with anti-CD3 and treating with ENTO or LANRA, singly and in combination with anti-PD1. CD3+ T-cell proliferation as measured by Ki67 expression and pSYK activation were assessed by quantitative single-cell immunofluorescence microscopy. Six formalin-fixed, paraffin-embedded BM biopsies from newly diagnosed AML pts were assessed for spatial expression of mRNA using a digital spatial profiling method. Results: In AML, mutations in NPM1 with co-expression of HOXA9/MEIS1 at baseline predicted anti-proliferative activity to ENTO in ex vivo drug sensitivity studies. Accordingly, treatment of leukemic cells with either ENTO or LANRA inhibited SYK auto-phosphorylation in a dose-dependent manner. Pathway analysis of archival RNA-seq data from pts enrolled in the Leukemia and Lymphoma Society BEAT AML Master Protocol (NCT03013998 [BAML-16-001-S6]) revealed gene expression signatures at baseline associated with the observed sensitivity to ENTO, including significant enrichment in the expression of genes associated with leukemogenesis, myeloid differentiation and immune regulation. This was supported by T-cell functional studies, which demonstrated that ENTO and LANRA, singly as well as in combination with anti-PD1, could restore T-cell proliferation in primary AML pt samples that exhibited suppression. Lastly, Cancer Transcriptome Atlas analysis on archival BM biopsies obtained from these AML pts showed a strong correlation between immune checkpoint response and gene expression associated with immune pathway signaling. Summary/Conclusion: Inhibition of SYK is a promising therapeutic approach in NPM1m AML. Our studies illustrate that ENTO and LANRA may restore T-cell proliferation in a subset of AML pts with dysfunctional T-cell responses, suggesting a novel mechanism of action. Additional studies are required to fully understand the mechanism of SYK-mediated antitumor immune responses in AML.