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1.
Blood ; 142(12): 1071-1081, 2023 09 21.
Artigo em Inglês | MEDLINE | ID: mdl-37294924

RESUMO

Rebalance of coagulation and anticoagulation to achieve a hemostatic effect has recently gained attention as an alternative therapeutic strategy for hemophilia. We engineered a humanized chimeric antibody, SR604, based on a previously published murine antibody, HAPC1573, which selectively blocks the anticoagulant activity of human activated protein C (APC). SR604 effectively blocked the anticoagulation activities of APC in human plasma deficient in various coagulation factors in vitro with affinities ∼60 times greater than that of HAPC1573. SR604 exhibited prophylactic and therapeutic efficacy in the tail-bleeding and knee-injury models of hemophilia A and B mice expressing human APC (humanized hemophilic mice). SR604 did not interfere with the cytoprotection and endothelial barrier function of APC, nor were there obvious toxicity effects in humanized hemophilic mice. Pharmacokinetic study showed a high bioavailability (106%) of subcutaneously injected SR604 in cynomolgus monkeys. These results demonstrate that SR604 is expected to be a safe and effective therapeutic and/or prophylactic agent with a prolonged half-life for patients with congenital factor deficiencies including hemophilia A and B.


Assuntos
Hemofilia A , Proteína C , Humanos , Camundongos , Animais , Proteína C/uso terapêutico , Hemofilia A/tratamento farmacológico , Modelos Animais de Doenças , Coagulação Sanguínea , Anticoagulantes/uso terapêutico
2.
FEBS Open Bio ; 13(7): 1253-1265, 2023 07.
Artigo em Inglês | MEDLINE | ID: mdl-37302810

RESUMO

Lymphocyte activation gene-3 (LAG-3) is a type I transmembrane protein with structural similarities to CD4. Overexpression of LAG-3 enables cancer cells to escape immune surveillance, while its blockade reinvigorates exhausted T cells and strengthens anti-infection immunity. Blockade of LAG-3 may have antitumor effects. Here, we generated a novel anti-LAG-3 chimeric antibody, 405B8H3(D-E), through hybridoma technology from monoclonal antibodies produced in mice. The heavy-chain variable region of the selected mouse antibody was grafted onto a human IgG4 scaffold, while a modified light-chain variable region was coupled to the human kappa light-chain constant region. 405B8H3(D-E) could effectively bind LAG-3-expressing HEK293 cells. Moreover, it could bind cynomolgus monkey (cyno) LAG-3 expressed on HEK293 cells with a higher affinity than the reference anti-LAG-3 antibody BMS-986016. Furthermore, 405B8H3(D-E) promoted interleukin-2 secretion and was able to block the interactions of LAG-3 with liver sinusoidal endothelial cell lectin and major histocompatibility complex II molecules. Finally, 405B8H3(D-E) combined with anti-mPD-1-antibody showed effective therapeutic potential in the MC38 tumor mouse model. Therefore, 405B8H3(D-E) is likely to be a promising candidate therapeutic antibody for immunotherapy.


Assuntos
Anticorpos Monoclonais Humanizados , Anticorpos Monoclonais , Camundongos , Humanos , Animais , Células HEK293 , Macaca fascicularis/metabolismo
3.
J Exp Med ; 219(9)2022 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-35881112

RESUMO

Disease relapse and treatment-induced immunotoxicity pose significant clinical challenges for patients with hematological cancers. Here, we reveal distinctive requirements for neutralizing TNF receptor ligands APRIL and BAFF and their receptor activity in MM and DLBCL, impacting protein translation and production in MM cells and modulating the translation efficiency of the ATM interactor (ATMIN/ACSIZ). Therapeutically, we investigated the use of BCMA decoy receptor (sBCMA-Fc) as an inhibitor of APRIL and BAFF. While wild-type sBCMA-Fc effectively blocked APRIL signaling in MM, it lacked activity in DLBCL due to its weak BAFF binding. To expand the therapeutic utility of sBCMA-Fc, we engineered an affinity-enhanced mutant sBCMA-Fc fusion molecule (sBCMA-Fc V3) 4- and 500-fold stronger in binding to APRIL and BAFF, respectively. The mutant sBCMA-Fc V3 clone significantly enhanced antitumor activity against both MM and DLBCL. Importantly, we also demonstrated an adequate toxicity profile and on-target mechanism of action in nonhuman primate studies.


Assuntos
Linfoma Difuso de Grandes Células B , Mieloma Múltiplo , Animais , Fator Ativador de Células B/genética , Antígeno de Maturação de Linfócitos B/genética , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/terapia , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/genética , Transdução de Sinais , Proteína Transmembrana Ativadora e Interagente do CAML , Membro 13 da Superfamília de Ligantes de Fatores de Necrose Tumoral/genética
4.
Clin Cancer Res ; 27(15): 4435-4448, 2021 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-34011561

RESUMO

PURPOSE: Ovarian cancer represents a major clinical hurdle for immune checkpoint blockade (ICB), with reported low patient response rates. We found that the immune checkpoint ligand PD-L2 is robustly expressed in patient samples of ovarian cancers and other malignancies exhibiting suboptimal response to ICB but not in cancers that are ICB sensitive. Therefore, we hypothesize that PD-L2 can facilitate immune escape from ICB through incomplete blockade of the PD-1 signaling pathway. EXPERIMENTAL DESIGN: We engineered a soluble form of the PD-1 receptor (sPD-1) capable of binding and neutralizing both PD-L2 and PD-L1 with ×200 and ×10,000 folds improvement in binding affinity over wild-type PD-1 leading to superior inhibition of ligand-mediated PD-1 activities. RESULTS: Both in vitro and in vivo analyses performed in this study demonstrated that the high-affinity sPD-1 molecule is superior at blocking both PD-L1- and PD-L2-mediated immune evasion and reducing tumor growth in immune-competent murine models of ovarian cancer. CONCLUSIONS: The data presented in this study provide justification for using a dual targeting, high-affinity sPD-1 receptor as an alternative to PD-1 or PD-L1 therapeutic antibodies for achieving superior therapeutic efficacy in cancers expressing both PD-L2 and PD-L1.


Assuntos
Inibidores de Checkpoint Imunológico/uso terapêutico , Neoplasias Ovarianas/tratamento farmacológico , Proteína 2 Ligante de Morte Celular Programada 1/antagonistas & inibidores , Animais , Resistencia a Medicamentos Antineoplásicos , Feminino , Humanos , Camundongos
5.
Sci Rep ; 9(1): 17830, 2019 11 28.
Artigo em Inglês | MEDLINE | ID: mdl-31780710

RESUMO

Programmed cell death 1 (PD-1) monoclonal antibodies have been approved by regulatory agencies for the treatment of various types of cancer, and the mechanism involves the restoration of T cell functions. We report herein the X-ray crystal structure of a fully human monoclonal antibody mAb059c fragment antigen-binding (Fab) in complex with the PD-1 extracellular domain (ECD) at a resolution of 1.70 Å. Structural analysis indicates 1) an epitope, comprising fragments from the C'D, BC and FG loops of PD-1, contributes to mAb059c interaction, 2) an unique conformation of the C'D loop and a different orientation of R86 enabling the capture of PD-1 by the antibody complementarity determining region (CDR) and the formation of one salt-bridge contact - ASP101(HCDR3):ARG86(PD-1), and 3) the contact of FG with light chain (LC) CDR3 is maintained by a second salt-bridge and two backbone hydrogen bonds. Interface analysis reveals that N-glycosylation sites 49, 74 and 116 on PD-1 do not contact mAb059c; while N58 in the BC loop is recognized by mAb059c heavy chain CDR1 and CDR2. Mutation of N58 attenuated mAb059c binding to PD-1. These findings and the novel anti-PD-1 antibody will facilitate better understanding of the mechanisms of the molecular recognition of PD-1 receptor by anti-PD-1 mAb and, thereby, enable the development of new therapeutics with an expanded spectrum of efficacy for unmet medical needs.


Assuntos
Adenocarcinoma/terapia , Anticorpos Monoclonais/química , Anticorpos Monoclonais/uso terapêutico , Antineoplásicos Imunológicos/química , Antineoplásicos Imunológicos/uso terapêutico , Neoplasias do Colo/terapia , Receptor de Morte Celular Programada 1/química , Animais , Afinidade de Anticorpos , Complexo Antígeno-Anticorpo/química , Linhagem Celular Tumoral , Modelos Animais de Doenças , Epitopos/química , Técnicas de Introdução de Genes , Células HEK293 , Humanos , Ligação de Hidrogênio , Fragmentos Fab das Imunoglobulinas/química , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptor de Morte Celular Programada 1/genética , Ligação Proteica , Domínios Proteicos , Transfecção
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