Editor’s Note: From December 9 to 12, 2023, the 65th American Society of Hematology (ASH) Annual Meeting was grandly held in San Diego, USA, in a hybrid format combining online and offline sessions. As one of the largest and most comprehensive international academic conferences in the field of hematology, experts and scholars from around the world gathered to explore the latest frontiers in hematologic diseases, new treatment methods and strategies, and disease management approaches. At this year’s ASH Annual Meeting, Professor Jin Jie and her team from the First Affiliated Hospital of Zhejiang University School of Medicine presented an oral report titled “IL1RAP-Specific T Cell Engager (TCE) Antibody Effectively Clears Leukemia Stem Cells (LSC) in Acute Myeloid Leukemia (AML).” “Tumor Watch” had the privilege of interviewing Professor Jin Jie on-site to discuss the research and its related content.
586 IL1RAP-Specific T Cell Engager (TCE) Antibody Efficiently Depletes Acute Myeloid Leukemia (AML) Leukemic Stem Cells (LSCs)
Research Introduction
LSCs are self-renewal primitive cells that initiate and maintain AML. Because they are highly therapy resistant, they are often responsible for treatment failures. Due to the lack of a specific antigen (Ag) and their quiescent nature, LSCs have been challenging to target and eliminate. IL1RAP is highly expressed on most human AML cell lines and primary blasts, including LSC-enriched CD34+CD38- cells, but not on normal hematopoietic stem cells (HSCs). To leverage this observation, we generated over 50 unique mouse IL1RAP monoclonal (m) antibodies (Abs), grouped them based on sequence, and selected representative members from the clades to generate murine-human chimeric mAbs. One of the mAbs, termed #24, had the strongest antibody-dependent cell-mediated cytotoxicity (ADCC) properties and the greatest in vivo anti-AML activity. Thus, it was advanced to generate a bispecific (Bs) anti-IL1RAP/CD3 TCE Ab termed BiF002, which incorporates a human IgG1 Fc mutated to lack FcR binding. Incubation of BIF002 with healthy donor T cells and AML cell lines or primary AML blasts induced IL1RAP dependent T cell activation, and Ab-dependent T cell lysis of leukemic cells in a dose-, time- and effector to target (E: T) ratio-dependent manner. The IC50 against MOLM13 and THP-1 was subnanomolar (0.047nM, 95%CI:0.036-0.060nM and 0.025nM, 95%CI:0.14-0.40nM, respectively), at E:T ratio 5:1 at 48 hours using resting T cells. The IC50 of IL1RAPpos AML bulk and CD34+ blasts (n=6) was also subnanomolar (0.45nM; 95%CI: 0.28-1.96nM). Co-cultures of normal CD34+ cells with BIF002 and T cells did not activate T cells or eliminate the normal CD34+ cells.
To test the antileukemic activity of BIF002 in vivo, we engrafted luciferase (luc)-expressing MOLM13 cells into NSG-SGM3 (NSGS) mice. On day (D) 3, in vitro expanded healthy donors’ T cells (3×106/mouse, weekly), with either BIF002 or a control IgG (10µg,) were administered intravenously (IV) every 3 days x 4-weeks. Mice treated with T cells + BIF002 exhibited a 100-fold greater tumor reduction on D33 than other groups, as measured by bioluminescence imaging (BLI), and had a significantly longer overall survival (OS; median 54.5 D) compared to IgG (27 D, p=0.0045), BIF002 alone (36 D, p=0.0049), or T cells + IgG (33 D, p=0.0062) treated controls. In initial dose finding studies using a similar schema, lower BIF002 doses (0.1 and 1 µg) were tested. Mice treated with T cells and 1 µg of BIF002 had a 100-fold tumor reduction on D29 and longer OS (median 57 D) compared to T cells + 0.1µg of BIF002 (31 D, p=0.0018) or T cells + vehicle (34 D, p=0.0019). We also engrafted luc-expressing primary blasts from a relapsed/refractory complex karyotype AML patient into NSGS mice that treated for 3 weeks using the above described schema (10µg). In this experiment, we used as control BIF026, a bispecific Ab (BsAb) carrying point mutations in both Fab arms to abrogate target binding to IL1RAP and CD3. Mice treated with T cells + BIF002 exhibited a >1000-fold reduction in disease burden and longer OS than BIF026 or BIF002 alone or T-cells+ BIF026-treated mice (median 46 D vs 26, 25 or 30 D, respectively, all comparisons, p=0.0003). In a second patient-derived xenograft (PDX) model, blasts from a relapsed complex karyotype AML patient were engrafted into NSG mice. Treatment was initiated on D10, with T cells given weekly along with 10µg of BIF002 or BIF026 for 3 weeks. After 70 days, all mice treated with T cells + BIF002 were alive, with no detectable blasts in peripheral blood or bone marrow (BM), while all vehicle (median 38 D, p=0.0002) and T cells + BIF026 (38 D, p=0.0001) treated controls had died of leukemia. To determine if T cells + BIF002 eradicated LSCs, we randomly selected 3 female mice from each group on D 34 and transplanted BM mononuclear cells (MNC) into secondary NSGS recipients (n=7). At D 90, no recipients of BM from donors treated with T cells + BIF002 had detectable blasts and remained alive, while all vehicle (median 26 D, p=0.0004) and T cells + BIF026 (median 26 D, p=0.0002) treated controls succumbed to AML. Importantly, no obvious hematopoietic or non-hematologic side effects were observed after BIF002 administration alone or with activated T cells in normal and leukemic mice.
In summary, we provide evidence of the subnanomolar potency and efficacy of BIF002, a novel anti-IL1RAP/CD3 TCE, in eliminating AML blasts, including LSCs. BIF002 is currently undergoing IND-enabling studies.
Expert Commentary
01
“Hematology Frontier”: First, could you please explain the expression of IL1RAP in AML cells and the significance of targeting IL1RAP in treatment?
Professor Jin Jie: IL1RAP is highly expressed in myeloid leukemia, accounting for 80% to 90% of cases. As it is expressed on the surface of AML tumor cells, it serves as an excellent target for antibody therapy.
02
“Hematology Frontier”: At this ASH conference, your team’s study (Abstract No: 586) focused on the dual-specificity antibody BiF002 targeting IL1RAP and CD3, showing significant activity in killing leukemia cells, including leukemia stem cells (LSCs). Could you provide insights into this research?
Professor Jin Jie: Besides high expression in myeloid leukemia, IL1RAP also exhibits elevated expression in leukemia stem cells, which is not present in normal stem cells. Therefore, antibodies based on IL1RAP can precisely target leukemia stem cells without affecting normal cells, unlike CD123 and CLL-1, which may have limited expression in normal stem cells. Through leukemia mouse models, especially patient-derived xenograft (PDX) models, we observed high expression of IL1RAP in leukemia stem cells. After antibody treatment, these stem cells were effectively eliminated. This indicates that IL1RAP has a significant advantage in targeting leukemia stem cells, making it a promising candidate for antibody or CAR-T therapy.
03
“Hematology Frontier”: Finally, what are your thoughts on the therapeutic efficacy and prospects of Chimeric Antigen Receptor T cells (CAR-T) targeting IL1RAP for AML? Could you discuss this in the context of related research?
Professor Jin Jie: We anticipate that IL1RAP will be a favorable CAR-T target because of its effective clearance of leukemia stem cells with minimal damage to normal stem cells, resulting in fewer side effects and off-target effects. We are currently conducting further research in this direction. In the future, we can develop dual-target antibodies by combining IL1RAP with CD3, providing an excellent immunotherapy for myeloid leukemia. Additionally, CAR-T therapy targeting IL1RAP holds great promise, especially in the context of relapsed or refractory leukemia, offering significant application prospects.
ASH 2023
Professor Jie Jin
Doctor of Medicine, Professor, Ph.D. Supervisor
Recipient of the Special Government Allowance from the State Council, Advanced Worker in the National Health System
Honorary Director of the Hematology Department, First Affiliated Hospital, Zhejiang University School of Medicine
Director of the Key Laboratory of Hematologic Malignancies (Diagnosis and Treatment), Zhejiang University
Leader of the Clinical Key Discipline of Hematology at Zhejiang University Hospital, National Health Commission
Director of the Clinical Medical Research Center for Hematologic Diseases in Zhejiang Province
Principal Investigator of Malignant Hematologic Diseases in the Cancer Research Institute, Zhejiang University
Leader of the Key Innovation Team in Leukemia Basic and Clinical Research in Zhejiang Province
Chairwoman of the Hematology Branch of the Chinese Medical Women’s Association
Former Chairwoman of the Hematologic Disease Translational Medicine Committee, Anti-Cancer Association
Executive Committee Member of the Hematology Society, Chinese Medical Association
Vice Executive Committee Member of the Integrated Hematology Society, Chinese Medical Association
Vice Executive Committee Member of the CSCO Anti-Leukemia Alliance
Executive Committee Member of the CSCO Anti-Lymphoma Alliance
Executive Committee Member of the Cross-Strait Hematology Society
Executive Committee Member of the Hematology Society, Chinese Health Promotion Association
Former Chairwoman of the Hematology Branch, Zhejiang Medical Association
President of the Hematology Branch, Zhejiang Provincial Medical Doctor Association
Published over 280 academic papers in SCI-indexed journals such as Lancet Oncology, Cell, Blood, Leukemia, and JHO. As the first awardee, received one National Science and Technology Progress Second Prize and two Zhejiang Provincial Science and Technology Progress First Prizes.
