Session: 614. Acute Lymphoblastic Leukemias: Therapies, Excluding Transplantation and Cellular Immunotherapies: Poster I
Hematology Disease Topics & Pathways:
Research, Lymphoid Leukemias, ALL, Translational Research, Combination therapy, Diseases, Therapies, Lymphoid Malignancies
Inhibition of T-cell activation and function by dasa or pona at 0.3 - 100 nM was tested in vitro over 3 days: (1) CD69 expression, (2) blina-mediated killing as specific lysis of CD19+ tumor cells (3) T-cell expansion analyzed by dye dilution or CD2+ fold change (FC), (4) cytokine secretion measured by cytokine catch or cytometric bead array.
Inhibition of TCR signaling was evaluated using a Jurkat-NFAT T-cell line stimulated for 6h with blina and CD19+ tumor cells. Phosphoflow of pAKT and pZAP70 was performed on CD3/CD28 activated primary human T cells. Toxicity of the TKIs against BCR-ABL+/- cell lines was evaluated after 3 days.
T-cell exhaustion upon continuous blina + dasa stimulation was assessed in an in vitro long-term culture system for continuous bispecific exposure (Philipp 2022). Functional assays as described in (1)-(4) were performed every 7 days with isolated T cells from long-term cultures.
Both TKIs inhibited T-cell function in a dose-dependent manner. Surprisingly, dasa was more potent than pona at low-intermediate doses in reducing T-cell function, shown by CD69 expression (mean % at TKI = 3 nM: basal = 70, dasa = 25, pona = 64, p<0.0001), cytotoxic function (mean % specific lysis at TKI = 12.5 nM: basal = 55, dasa = 0, pona = 54, p<0.0001), T-cell proliferation (mean % proliferated at TKI = 1.5 nM: basal = 62, dasa = 25, pona = 51, p<0.0001), as well as IFN-𝛾 secretion (normalized mean fluorescence intensity (MFI) at TKI = 12.5 nM: basal = 655, dasa = 0, pona = 924, p<0.0001). We also observed that NFAT engagement was significantly lower in dasa-treated Jurkat-NFAT cells (relative luminescence units at TKI 12.5 nM: basal = 1620, dasa = 392, pona = 996, p<0.001). Similarly, phosphoflow analyses revealed lower levels of pAKT and pZAP70 upon CD3/CD28 + dasa versus pona treatment (MFI of pAKT at TKI = 25 nM: basal = 2347, dasa = 1642, pona = 2045). Importantly, both TKIs induced cell death of BCR-ABL+ but not BCR-ABL- cell lines at concentrations >3 nM.
Interestingly, 14 days continuously stimulated T cells with blina + dasa (12.5 nM) did not lead to co-expression of inhibitory checkpoints (mean % of PD-1+Tim-3+LAG-3+ T cells on day 14: blina = 31, +dasa = 0.6, p<0.01). Furthermore, when isolated from long-term cultures and tested in functional assays, blina + dasa treated T cells maintained high cytokine secretion (mean IL-2 in pg/ml on day 14: blina = 730, blina + dasa = 5638, p<0.01), granzyme B expression (mean MFI ratio of CD8+ on day 14: blina = 96, blina + dasa = 335, p<0.01) and cytotoxic activity (mean % specific lysis on day 14: blina = 61, blina + dasa = 80).
Together, these findings indicate that while both TKIs confer comparable toxicity to BCR-ABL+ leukemia cells, dasa dampens TCR signaling and T-cell effector function more potently than pona. Strikingly, dasa can ameliorate T-cell exhaustion upon continuous blina exposure. Our data suggest that low-dose TKIs can fine-tune T-cell activity and might play a considerable role for the clinical success of combinatorial bispecific and TKI therapy in Ph+ BCP-ALL.
Disclosures: Buecklein: Gilead/Kite: Other: Travel Funding, Research Funding; BMS: Research Funding; Miltenyi Biotech: Research Funding; Roche: Honoraria, Research Funding; Pierre Fabre: Other: Travel Funding; Priothera: Consultancy; Pfizer: Consultancy, Honoraria, Speakers Bureau; Amgen: Consultancy. Subklewe: Seagen: Research Funding; Ichnos Sciences: Consultancy, Honoraria; Roche: Consultancy, Honoraria, Other: Travel Support, Research Funding, Speakers Bureau; Pfizer: Consultancy, Honoraria, Other: Travel Support, Speakers Bureau; AstraZeneca: Speakers Bureau; Takeda: Consultancy, Honoraria, Research Funding; BMS/Celgene: Consultancy, Honoraria, Research Funding, Speakers Bureau; Janssen: Consultancy, Honoraria, Research Funding, Speakers Bureau; Amgen: Consultancy, Honoraria, Research Funding; Miltenyi Biotec: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding, Speakers Bureau; Gilead/Kite: Consultancy, Honoraria, Other: Travel Support, Research Funding, Speakers Bureau; AvenCell: Consultancy, Honoraria; Incyte Biosciences: Consultancy, Honoraria; Molecular Partners: Consultancy, Honoraria, Research Funding; GSK: Speakers Bureau; LAWG: Speakers Bureau; Springer Healthcare: Speakers Bureau; AbbVie: Consultancy, Honoraria; Autolus: Consultancy, Honoraria; advesya (CanCell Therapeutics): Consultancy, Honoraria; Genmab US: Consultancy, Honoraria; Interius BioTherapeutics: Consultancy, Honoraria; Nektar Therapeutics: Consultancy, Honoraria; Orbital Therapeutics: Consultancy, Honoraria; Sanofi: Consultancy, Honoraria; Scare: Consultancy, Honoraria.