Session: 641. Chronic Lymphocytic Leukemia: Basic and Translational: Poster II
Hematology Disease Topics & Pathways:
Research, Lymphoid Leukemias, Adult, Translational Research, CLL, Elderly, Diseases, Immunology, Lymphoid Malignancies, Biological Processes, Molecular biology, Study Population, Human
Peripheral blood (PB) from 73 treatment-naïve CLL patients was collected and analyzed by flow cytometry. Median age was 65 years and 40% had unmutated IGHV. A subset of circulating CLL cells that expressed PD-1 was detected in all patients. The percentage of circulating PD-1+ CLL cells was variable and ranged from 10% to 85% (mean 43%). Importantly, nearly all the CLL cells that expressed the proliferation marker Ki-67 also expressed PD-1 (median >96%). PD-1+ CLL cells also had higher expression of CD5 and lower expression of CXCR4 (MFI fold change 1.49 and 0.49, respectively, p<0.0001), in line with recently divided status. RNA sequencing analysis on flow-sorted cells with a PD-1 MFI >90th percentile (PD-1hi) and PD-1- cells revealed 6580 differentially expressed genes (FDR 0.05). These genes were associated with B-cell activation, proliferation, migration, signaling, apoptosis, and metabolism. Notably, pathways associated with cell activation, cell cycle, oxidative phosphorylation, and B-cell receptor (BCR), toll-like receptor (TLR), and PD-1 signaling were all upregulated in PD-1hi cells.
We hypothesized that PD-1 expression in CLL cells might be induced through BCR and TLR signaling. To test this, we stimulated CLL cells ex vivo either through the BCR using anti-IgM antibodies, or through TLR9 with CpG and quantified PD-1 expression over time. The percentage of PD-1+ cells significantly increased within 24 hours after stimulation with either agent. Notably, concomitant inhibition of BTK-dependent signaling by ibrutinib resulted in near-complete abrogation of the effect of anti-IgM, and a marked decrease in CPG’s effect on PD-1 expression.
To confirm the effect of BTKi on PD-1 expression in vivo, we analyzed the PB of 40 patients receiving either frontline or later line ibrutinib or acalabrutinib therapy for ≥1 month (median age: 65 years, 55% unmutated IGHV). In patients adherent to therapy, PD-1 was expressed in <10% of remaining circulating CLL cells. The percentage of circulating PD-1+ cells decreased in all patients within 2 weeks after treatment initiation and remained low while on therapy. Importantly, 13-90% of circulating CLL cells expressed PD-1 in all 18 patients who were found to have clinical progression from BTKi therapy within 3 months of sample acquisition (median age: 66 years, 44% unmutated IGHV). Circulating PD-1+ cells that re-emerged at the time of BTKi progression overexpressed genes associated with cell activation, cell cycle, oxidative phosphorylation, cytokine response, and PD-1, BCR, and TLR signaling. Additional clinical characteristics and biomarkers for all cohorts will be reported.
Overall, our data indicate that PD-1 identifies a subset of circulating CLL cells that are transcriptionally more active and have recently undergone proliferation. We showed that PD-1 expression is induced via BCR and TLR signaling through BTK, implying that PD-1 could be a marker of CLL cell activation. Moreover, increased circulating PD-1+ CLL cells in patients on BTKi therapy could serve as an early marker of BTKi resistance.
Disclosures: Chang: AstraZeneca: Consultancy, Honoraria. Koff: Viracta Therapeutics: Research Funding; AbbVie: Consultancy; BeiGene: Consultancy. Flowers: Karyopharm: Consultancy; Pharmacyclics / Janssen: Consultancy; Iovance: Research Funding; Adaptimmune: Research Funding; Bristol Myers Squibb: Consultancy; Janssen Pharmaceuticals: Research Funding; Genmab: Consultancy; Takeda: Research Funding; Burroughs Wellcome Fund: Research Funding; Allogene: Research Funding; Pharmacyclics: Research Funding; Pfizer: Research Funding; Eastern Cooperative Oncology Group: Research Funding; 4D: Research Funding; Novartis: Research Funding; EMD Serono: Research Funding; Bio Ascend: Consultancy; TG Therapeutics: Research Funding; Acerta: Research Funding; Foresight Diagnostics: Consultancy, Current holder of stock options in a privately-held company; Xencor: Research Funding; Cellectis: Research Funding; Gilead: Consultancy, Research Funding; Seagen: Consultancy; N-Power Medicine: Consultancy, Current holder of stock options in a privately-held company; Sanofi: Research Funding; BostonGene: Research Funding; Ziopharm National Cancer Institute: Research Funding; Amgen: Research Funding; Nektar: Research Funding; Morphosys: Research Funding; Cancer Prevention and Research Institute of Texas: CPRIT Scholar in Cancer Research: Research Funding; Kite: Research Funding; Denovo Biopharma: Consultancy; Celgene: Consultancy, Research Funding; BeiGene: Consultancy; Bayer: Consultancy, Research Funding; AstraZeneca: Consultancy; Genentech/Roche: Consultancy, Research Funding; Guardant: Research Funding; Spectrum: Consultancy; AbbVie: Consultancy, Research Funding.
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