HUMANIZATION AND FUNCTIONAL CHARACTERIZATION OF A MONOCLONAL ANTIBODY AGAINST THE INTERLEUKIN-7 RECEPTOR ALPHA CHAIN IN ACUTE LYMPHOBLASTIC LEUKEMIA
- 1Universidade Estadual de Campinas (UNICAMP), Campinas, SP, Brazil; Centro de Pesquisa Boldrini (CPB), Campinas, SP, Brazil
- 2Universidade de Brasília (UnB), Brasília, DF, Brazil
Abstract
Background: Therapeutic monoclonal antibodies are promising tools for the treatment of acute lymphoblastic leukemia (ALL). Our group previously developed a murine anti-IL-7Rα antibody (mAb#1) capable of recognizing IL-7Rα on leukemic cells. However, murine antibodies may induce immunogenic responses, limiting their clinical use and supporting the need for humanization. Aim: To generate and characterize a humanized version of mAb#1, termed HumAb#1, using CDR grafting, and to evaluate its structural conservation, antigen recognition, effects on intracellular signaling, and in vivo antileukemic activity. Materials and methods: Human germline sequences were selected as acceptor frameworks, and the CDRs of mAb#1 were grafted onto these sequences. Strategic backmutations were introduced to preserve structural and functional features related to antigen recognition. The humanized heavy and light chains were expressed in ExpiCHO-S cells. Structural conservation was assessed by modeling and alignment. Antigen recognition was evaluated by dot blot, flow cytometry, and binding assays using recombinant IL-7Rα. The effects of HumAb#1 on IL-7-induced signaling were analyzed by Western blot for phosphorylated STAT5, ERK1/2, and AKT. Antileukemic activity was evaluated in NSG mice xenografted with RS4;11 cells. Results: HumAb#1 showed high structural conservation relative to the parental antibody, with an RMSD of 0.422 Å. Flow cytometry demonstrated selective recognition of IL-7Rα-positive cells, with no detectable binding to negative cells. Assays using the recombinant receptor confirmed preservation of antigen recognition. HumAb#1 enhanced IL-7-induced signaling, with increased phosphorylation of STAT5, ERK1/2, and AKT. Despite this in vitro effect, treatment with HumAb#1 reduced leukemia burden in NSG mice xenografted with RS4;11 cells, as assessed by bioluminescence imaging. Discussion: Preservation of antigen recognition after humanization indicates that the CDR-grafting and backmutation strategy maintained important structural features of the parental antibody. The increased IL-7-induced phosphorylation of STAT5, ERK1/2, and AKT observed in vitro, together with the reduction in leukemia burden in vivo, suggests that the antileukemic activity of HumAb#1 may involve mechanisms beyond direct inhibition of IL-7Rα signaling. This apparent dissociation between signaling modulation and therapeutic activity highlights the need for further investigation of its mechanism of action. Conclusion: HumAb#1 retained structural and antigen-recognition properties of the parental antibody and demonstrated in vivo antileukemic activity, supporting further mechanistic and preclinical investigation.
Related
- Efficacy and safety of XKH004, a novel dual interleukin-17A/F inhibitory monoclonal antibody, in active ankylosing spondylitis: A 52-week phase 3 randomized study
- Characterization of monoclonal and patient-derived antiplatelet factor 4 antibodies in platelet factor 4 and platelet factor 4/polyanion chemiluminescence assays
- COMPREHENSIVE CLINICAL EVALUATION OF THE EXENT ANALYZER FOR DETECTION AND CHARACTERIZATION OF MONOCLONAL GAMMOPATHIES
- Development and Preliminary Psychometric Evaluation of a Questionnaire on Comprehensive Protection, Quality, and Humanization in Pediatric Hospital Care
- Structural and Functional Characterization of a Novel Broad-Spectrum Oligosaccharide Uptake Channel OmpV25 in Vibrio campbellii
BibTeX
@article{fonseca2026humanization,
title = {{HUMANIZATION AND FUNCTIONAL CHARACTERIZATION OF A MONOCLONAL ANTIBODY AGAINST THE INTERLEUKIN-7 RECEPTOR ALPHA CHAIN IN ACUTE LYMPHOBLASTIC LEUKEMIA}},
author = {Joey Ramone Ferreira FONSECA and Fernando Mesquita de Sousa de LIMA and João Mateus Silva FEITOZA and Aryanny Paula Sousa FERREIRA and Marcelo de Macedo BRIGIDO and Priscila Pini ZENATTI},
journal = {Hematology, Transfusion and Cell Therapy},
year = {2026},
month = oct,
doi = {10.1016/j.htct.2026.107599},
url = {https://doi.org/10.1016/j.htct.2026.107599},
note = {CC BY},
}
RIS
TY - JOUR
TI - HUMANIZATION AND FUNCTIONAL CHARACTERIZATION OF A MONOCLONAL ANTIBODY AGAINST THE INTERLEUKIN-7 RECEPTOR ALPHA CHAIN IN ACUTE LYMPHOBLASTIC LEUKEMIA
AU - Joey Ramone Ferreira FONSECA
AU - Fernando Mesquita de Sousa de LIMA
AU - João Mateus Silva FEITOZA
AU - Aryanny Paula Sousa FERREIRA
AU - Marcelo de Macedo BRIGIDO
AU - Priscila Pini ZENATTI
PY - 2026
DA - 2026/10/23
JO - Hematology, Transfusion and Cell Therapy
DO - 10.1016/j.htct.2026.107599
UR - https://doi.org/10.1016/j.htct.2026.107599
AB - Background: Therapeutic monoclonal antibodies are promising tools for the treatment of acute lymphoblastic leukemia (ALL). Our group previously developed a murine anti-IL-7Rα antibody (mAb#1) capable of recognizing IL-7Rα on leukemic cells. However, murine antibodies may induce immunogenic responses, limiting their clinical use and supporting the need for humanization. Aim: To generate and characterize a humanized version of mAb#1, termed HumAb#1, using CDR grafting, and to evaluate its structural conservation, antigen recognition, effects on intracellular signaling, and in vivo antileukemic activity. Materials and methods: Human germline sequences were selected as acceptor frameworks, and the CDRs of mAb#1 were grafted onto these sequences. Strategic backmutations were introduced to preserve structural and functional features related to antigen recognition. The humanized heavy and light chains were expressed in ExpiCHO-S cells. Structural conservation was assessed by modeling and alignment. Antigen recognition was evaluated by dot blot, flow cytometry, and binding assays using recombinant IL-7Rα. The effects of HumAb#1 on IL-7-induced signaling were analyzed by Western blot for phosphorylated STAT5, ERK1/2, and AKT. Antileukemic activity was evaluated in NSG mice xenografted with RS4;11 cells. Results: HumAb#1 showed high structural conservation relative to the parental antibody, with an RMSD of 0.422 Å. Flow cytometry demonstrated selective recognition of IL-7Rα-positive cells, with no detectable binding to negative cells. Assays using the recombinant receptor confirmed preservation of antigen recognition. HumAb#1 enhanced IL-7-induced signaling, with increased phosphorylation of STAT5, ERK1/2, and AKT. Despite this in vitro effect, treatment with HumAb#1 reduced leukemia burden in NSG mice xenografted with RS4;11 cells, as assessed by bioluminescence imaging. Discussion: Preservation of antigen recognition after humanization indicates that the CDR-grafting and backmutation strategy maintained important structural features of the parental antibody. The increased IL-7-induced phosphorylation of STAT5, ERK1/2, and AKT observed in vitro, together with the reduction in leukemia burden in vivo, suggests that the antileukemic activity of HumAb#1 may involve mechanisms beyond direct inhibition of IL-7Rα signaling. This apparent dissociation between signaling modulation and therapeutic activity highlights the need for further investigation of its mechanism of action. Conclusion: HumAb#1 retained structural and antigen-recognition properties of the parental antibody and demonstrated in vivo antileukemic activity, supporting further mechanistic and preclinical investigation.
SN - 2531-1379
SN - 2531-1387
ER -