One of the G protein-coupled receptors, C-C chemokine receptor 5 (CCR5), is an important regulator for the activation of T and B lymphocytes, dendritic cells, natural killer cells, and macrophages. Upon binding to its ligands, CCR5 activates downstream signaling, which is an important regulator in the innate and adaptive immune response through the promotion of lymphocyte migration and the secretion of proinflammatory cytokines. Anti-CCR5 monoclonal antibodies (mAbs) have been developed and evaluated in clinical trials for tumors and inflammatory diseases. In this study, we developed novel mAbs for mouse CCR5 (mCCR5) using the N-terminal peptide immunization. Among the established anti-mCCR5 mAbs, C5Mab-4 (rat IgG2a, kappa) and C5Mab-8 (rat IgG1, kappa), recognized mCCR5-overexpressing Chinese hamster ovary-K1 (CHO/mCCR5) and an endogenously mCCR5-expressing cell line (L1210) by flow cytometry. The dissociation constant (KD) values of C5Mab-4 and C5Mab-8 for CHO/mCCR5 were determined as 3.5 × 10−8 M and 7.3 × 10−9 M, respectively. Furthermore, both C5Mab-4 and C5Mab-8 could detect mCCR5 by western blotting. These results indicated that C5Mab-4 and C5Mab-8 are useful for detecting mCCR5 by flow cytometry and western blotting and provide a possibility to obtain the proof of concept in preclinical studies.
MohamedH, GurrolaT, BermanR, et al.Targeting CCR5 as a Component of an HIV-1 Therapeutic Strategy. Front Immunol, 2021; 12:816515; doi: 10.3389/fimmu.2021.816515
3.
ZengZ, LanT, WeiY, et al.CCL5/CCR5 axis in human diseases and related treatments. Genes Dis, 2022; 9(1):12–27; doi: 10.1016/j.gendis.2021.08.004
4.
HamidR, AlazizM, MahalAS, et al.The role and therapeutic targeting of CCR5 in Breast Cancer. Cells, 2023; 12(18); doi: 10.3390/cells12182237
5.
AldinucciD, CasagrandeN. Inhibition of the CCL5/CCR5 Axis against the Progression of Gastric Cancer. Int J Mol Sci, 2018; 19(5); doi: 10.3390/ijms19051477
6.
AlkhatibG. The biology of CCR5 and CXCR4. Curr Opin HIV AIDS, 2009; 4(2):96–103; doi: 10.1097/COH.0b013e328324bbec
7.
Martin-BlondelG, BrassatD, BauerJ, et al.CCR5 blockade for neuroinflammatory diseases–beyond control of HIV. Nat Rev Neurol, 2016; 12(2):95–105; doi: 10.1038/nrneurol.2015.248
8.
ChuaRL, LukassenS, TrumpS, et al.COVID-19 severity correlates with airway epithelium-immune cell interactions identified by single-cell analysis. Nat Biotechnol, 2020; 38(8):970–979; doi: 10.1038/s41587-020-0602-4
9.
WoodA, ArmourD. The discovery of the CCR5 receptor antagonist, UK-427,857, a new agent for the treatment of HIV infection and AIDS. Prog Med Chem, 2005; 43:239–271; doi: 10.1016/s0079-6468(05)43007-6
10.
ScurciI, MartinsE, HartleyO. CCR5: Established paradigms and new frontiers for a ‘celebrity’ chemokine receptor. Cytokine, 2018; 109:81–93; doi: 10.1016/j.cyto.2018.02.018
11.
QiB, FangQ, LiuS, et al.Advances of CCR5 antagonists: From small molecules to macromolecules. Eur J Med Chem, 2020; 208:112819; doi: 10.1016/j.ejmech.2020.112819
12.
OuchidaT, IsodaY, NakamuraT, et al.Establishment of a novel anti-mouse CCR1 monoclonal antibody C(1)Mab-6. Monoclon Antib Immunodiagn Immunother, 2024; 43(2):67–74; doi: 10.1089/mab.2023.0032
13.
TateyamaN, AsanoT, SuzukiH, et al.Epitope mapping of anti-mouse CCR3 monoclonal antibodies using flow cytometry. Antibodies (Basel), 2022; 11(4); doi: 10.3390/antib11040075
14.
SaitoM, HarigaeY, LiG, et al.C(3)Mab-2: An anti-mouse CCR3 monoclonal antibody for immunocytochemistry. Monoclon Antib Immunodiagn Immunother, 2022; 41(1):45–49; doi: 10.1089/mab.2021.0050
15.
AsanoT, SuzukiH, TanakaT, et al.C(3)Mab-3: A monoclonal antibody for mouse CC chemokine receptor 3 for flow cytometry. Monoclon Antib Immunodiagn Immunother, 2022; 41(2):74–79; doi: 10.1089/mab.2021.0062
16.
SuzukiH, TanakaT, LiG, et al.Development of a sensitive anti-mouse CCR5 monoclonal antibody for flow cytometry. Monoclon Antib Immunodiagn Immunother, 2024; doi: 10.1089/mab2024.0004
17.
SuzukiH, SaitoM, AsanoT, et al.C(8)Mab-3: An anti-mouse CCR8 monoclonal antibody for immunocytochemistry. Monoclon Antib Immunodiagn Immunother, 2022; 41(2):110–114; doi: 10.1089/mab.2022.0002
18.
SaitoM, TanakaT, AsanoT, et al.C(8)Mab-2: An anti-mouse C-C motif chemokine receptor 8 monoclonal antibody for immunocytochemistry. Monoclon Antib Immunodiagn Immunother, 2022; 41(2):115–119; doi: 10.1089/mab.2021.0045
19.
SaitoM, SuzukiH, TanakaT, et al.Development of an anti-mouse CCR8 monoclonal antibody (C(8)Mab-1) for flow cytometry and immunocytochemistry. Monoclon Antib Immunodiagn Immunother, 2022; 41(6):333–338; doi: 10.1089/mab.2021.0069
20.
LiG, TanakaT, SuzukiH, et al.Cx1Mab-1: A novel anti-mouse CXCR1 monoclonal antibody for flow cytometry. Monoclon Antib Immunodiagn Immunother. 2024; 43(2):59–66; doi: 10.1089/mab.2023.0031
21.
OuchidaT, IsodaY, TanakaT, et al.Cx(3)Mab-4: A novel anti-mouse CXCR3 monoclonal antibody for flow cytometry. Monoclon Antib Immunodiagn Immunother, 2024; doi: 10.1089/mab.2023.0024
22.
OuchidaT, SuzukiH, TanakaT, et al.Cx(4)Mab-1: A novel anti-mouse CXCR4 monoclonal antibody for flow cytometry. Monoclon Antib Immunodiagn Immunother, 2023; 43(1):10–16; doi: 10.1089/mab.2023.0023
23.
TanakaT, LiG, AsanoT, et al.Development of a novel anti-mouse CCR2 monoclonal antibody (C(2)Mab-6) by N-terminal peptide immunization. Monoclon Antib Immunodiagn Immunother, 2022; 41(2):80–86; doi: 10.1089/mab.2021.0063
24.
AsanoT, SuzukiH, GotoN, et al.Establishment of novel anti-mouse CCR3 monoclonal antibodies (C(3)Mab-6 and C(3)Mab-7) by N-terminal peptide immunization. Monoclon Antib Immunodiagn Immunother, 2022; 41(2):94–100; doi: 10.1089/mab.2021.0065
25.
TakeiJ, SuzukiH, AsanoT, et al.Development of a novel anti-mouse CCR4 monoclonal antibody (C(4)Mab-1) by N-Terminal peptide immunization. Monoclon Antib Immunodiagn Immunother, 2022; 41(2):87–93; doi: 10.1089/mab.2021.0064
26.
KobayashiH, AsanoT, SuzukiH, et al.Establishment of a sensitive monoclonal antibody against mouse CCR9 (C(9)Mab-24) for flow cytometry. Monoclon Antib Immunodiagn Immunother, 2023; 42(1):15–21; doi: 10.1089/mab.2022.0032
27.
KitamuraK, SuzukiH, KanekoMK, et al.Cx(6)Mab-1: A novel anti-mouse CXCR6 monoclonal antibody established by n-terminal peptide immunization. Monoclon Antib Immunodiagn Immunother, 2022; 41(3):133–141; doi: 10.1089/mab.2022.0010
28.
KatoY, KanekoMK, KunoA, et al.Inhibition of tumor cell-induced platelet aggregation using a novel anti-podoplanin antibody reacting with its platelet-aggregation-stimulating domain. Biochem Biophys Res Commun, 2006; 349(4):1301–1307; doi: 10.1016/j.bbrc.2006.08.171
29.
KatoY. Specific monoclonal antibodies against IDH1/2 mutations as diagnostic tools for gliomas. Brain Tumor Pathol, 2015; 32(1):3–11; doi: 10.1007/s10014-014-0202-4
30.
IkotaH, NobusawaS, AraiH, et al.Evaluation of IDH1 status in diffusely infiltrating gliomas by immunohistochemistry using anti-mutant and wild type IDH1 antibodies. Brain Tumor Pathol, 2015; 32(4):237–244; doi: 10.1007/s10014-015-0222-8
31.
TsuiCK, GuptaA, BassikMC. Finding host targets for HIV therapy. Nat Genet, 2017; 49(2):175–176; doi: 10.1038/ng.3777
32.
FarzanM, MirzabekovT, KolchinskyP, et al.Tyrosine sulfation of the amino terminus of CCR5 facilitates HIV-1 entry. Cell, 1999; 96(5):667–676; doi: 10.1016/s0092-8674(00)80577-2
33.
LudemanJP, StoneMJ. The structural role of receptor tyrosine sulfation in chemokine recognition. Br J Pharmacol, 2014; 171(5):1167–1179; doi: 10.1111/bph.12455
34.
IsodaY, TanakaT, SuzukiH, et al.Epitope mapping of an anti-mouse CXCR6 monoclonal antibody (Cx(6)Mab-1) using the 2 × Alanine Scanning Method. Monoclon Antib Immunodiagn Immunother, 2022; 41(5):275–278; doi: 10.1089/mab.2022.0019
35.
IsaikinaP, TsaiCJ, DietzN, et al.Structural basis of the activation of the CC chemokine receptor 5 by a chemokine agonist. Sci Adv, 2021; 7(25); doi: 10.1126/sciadv.abg8685
36.
TanQ, ZhuY, LiJ, et al.Structure of the CCR5 chemokine receptor-HIV entry inhibitor maraviroc complex. Science, 2013; 341(6152):1387–1390; doi: 10.1126/science.1241475
37.
AbayevM, RodriguesJ, SrivastavaG, et al.The solution structure of monomeric CCL5 in complex with a doubly sulfated N-terminal segment of CCR5. Febs J, 2018; 285(11):1988–2003; doi: 10.1111/febs.14460
38.
SunD, SunY, JanezicE, et al.Structural basis of antibody inhibition and chemokine activation of the human CC chemokine receptor 8. Nat Commun, 2023; 14(1):7940; doi: 10.1038/s41467-023-43601-8
39.
ZhouM, GreenhillS, HuangS, et al.CCR5 is a suppressor for cortical plasticity and hippocampal learning and memory. Elife, 2016; 5:5; doi: 10.7554/eLife.20985
40.
ShenY, ZhouM, CaiD, et al.CCR5 closes the temporal window for memory linking. Nature, 2022; 606(7912):146–152; doi: 10.1038/s41586-022-04783-1
41.
HeidebrinkJL, PaulsonHL. Lessons learned from approval of aducanumab for Alzheimer’s Disease. Annu Rev Med, 2024; 75:99–111; doi: 10.1146/annurev-med-051022-043645
42.
HuangHY, SalinasS, CornellJ, et al.CCR5 regulates Aβ(1–42)-induced learning and memory deficits in mice. Neurobiol Learn Mem, 2024; 208:107890; doi: 10.1016/j.nlm.2024.107890
Supplementary Material
Please find the following supplemental material available below.
For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.
For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.
