Session: 614. Acute Lymphoblastic Leukemias: Biomarkers, Molecular Markers, and Minimal Residual Disease in Diagnosis and Prognosis: Poster III
Hematology Disease Topics & Pathways:
Research, Fundamental Science, Lymphoid Leukemias, ALL, Translational Research, Diseases, Immunology, Lymphoid Malignancies, Biological Processes
Methods: We created a novel mouse strain, HV1, that expresses a fixed T-cell receptor with specificity for a leukemia neoantigenic peptide derived from the BCR-ABL oncogene. Naïve HV1 CD4+ T-cells (10,000/mouse) were adoptively transferred into congenically distinct naïve recipients, followed 24h later by challenge with a BCR-ABL+ cell line (“LM138”; 2,500 cells/mouse). scATACseq, scRNAseq, multiplex immunofluorescence (Cellscape; Canopy-Biosciences) and functional studies were used to analyze HV1 cells during leukemia progression or treatment. To model MRD+ remissions, mice were treated with a 5-day course of nilotinib (75 mg/kg po). Nilotinib was then combined with anti-PDL1 blockade (10 mg/kg i.p.) to model curative therapy. Observations from adoptive transfer experiments were compared to bone marrow aspirates of patients with newly diagnosed with BCR-ABL+ or BCR-ABL- B-ALL. Survival analysis using log-rank testing was applied to outcomes from the TARGET database.
Results: UMAp clustering of HV1 cells at day 9 of leukemia challenge confirmed a canonical bifurcation into early T-follicular helper (Tfh) and Th1 subsets. By a late leukemia timepoint (day 17), both Tfh- and Th1-polarized HV1s adopted the epigenetic, transcriptomic, and phenotypic states (FOXP3-/IL10+) of Type-1 regulatory cells (Tr1s). Tr1-differentiated HV1s suppressed bystander CD8+ T-cells in co-culture assays. Leukemia cells co-localized with CD4+ T-cells but not CD8+ T-cells, and were found in closer proximity, on average, to CD4+ rather than CD8+ T-cells (mean distance 117 v. 143 um, p <0.001, student T-test). Ki67- LM138s preferentially maintained MHCII expression. HV1 subset proportions were significantly altered after treatment with nilotinib + anti-PDL1 therapy, resulting in a lower frequency of Tr1 cells and a higher frequency of CX3CR1+/AHNAKhi eff/mem cells, as compared to nilotinib only. CD8+ T-cell subset distributions were unchanged. We confirmed that similar CD4+ T-cell subsets are observable in scRNAseq datasets from patient samples. We interrogated the TARGET B-ALL database, using a Tr1 gene signature; patients with samples in the highest quartile of Tr1 signature expression experienced inferior overall survival as compared to those with the lowest (p =0.02, log-rank test).
Conclusions: Here, we show that neoantigen-specific CD4+ T-cells spontaneously develop into PD1+/TIM3+ Tr1s in the leukemia microenvironment. The relative proximity of CD4+ T-cells to LM138 cells and selective preservation of MHCII positivity in Ki67- LM138 cells implies that neoantigen-specific CD4+ T-cells may adopt Tr1 states after direct instruction by quiescent leukemia cells. This agrees with recent findings that neoantigen-specific CD4+ T-cells develop into Tr1s after direct contact with mutation-harboring hematopoetic stem cells (HSCs). Thus, Tr1s protect residual leukemia cells from immune attack by maladaptively recapitulating an immune circuit of the HSC niche. Eradication of residual B-ALL occurred concurrent with therapeutic reprogramming of neoantigen-specific CD4+ T-cells from Tr1 towards unique CX3CR1+/AHNAKhi mem/eff differentiation states.
Disclosures: Springer: Bruker Spatial Biology: Current Employment. Meshik: Bruker Spatial Biology: Current Employment. Bachanova: Citius: Research Funding; CRISPR: Consultancy; Astra Zeneca: Consultancy; Allogene: Consultancy; Miltenyi: Other: DSMB; Incyte: Research Funding; Gamida Cell: Research Funding.