Diversity in the HLA-I Recognition of HLA-F Monoclonal Antibodies: HLA-F or HLA-Ib Monospecific, HLA-E or HLA-G Bispecific Antibodies with or without HLA-Ia Reactivity

Mepur H Ravindranath; Narendranath M Ravindranath; Carly J Amato-Menker; Fatiha El Hilali; Edward J Filippone

doi:10.3390/antib13010008

Diversity in the HLA-I Recognition of HLA-F Monoclonal Antibodies: HLA-F or HLA-Ib Monospecific, HLA-E or HLA-G Bispecific Antibodies with or without HLA-Ia Reactivity

Antibodies (Basel). 2024 Jan 31;13(1):8. doi: 10.3390/antib13010008.

Authors

Mepur H Ravindranath^{1

2}, Narendranath M Ravindranath³, Carly J Amato-Menker⁴, Fatiha El Hilali⁵, Edward J Filippone⁶

Affiliations

¹ Department of Hematology and Oncology, Children's Hospital, Los Angeles, CA 90027, USA.
² Terasaki Foundation Laboratory, Santa Monica, CA 90064, USA.
³ Norris Dental Science Center, Herman Ostrow School of Dentistry, University of Southern California, Los Angeles, CA 90089, USA.
⁴ Department of Microbiology, Immunology and Cell Biology, School of Medicine, West Virginia University, Morgantown, WV 26506, USA.
⁵ Medico-Surgical, Biomedicine and Infectiology Research Laboratory, The Faculty of Medicine and Pharmacy of Laayoune & Agadir, Ibnou Zohr University, Agadir 80000, Morocco.
⁶ Division of Nephrology, Department of Medicine, Sidney Kimmel Medical College, Thomas Jefferson University, Philadelphia, PA 19145, USA.

Abstract

Previous investigators have used various anti-HLA-F monoclonal antibodies (mAbs) to demonstrate that the tissue distribution of HLA-F is highly restricted. Notably, these mAbs differed in their immunodiagnostic capabilities. Specifically, mAbs Fpep1.1 and FG1 detected HLA-F intracellularly in B cells but not on the cell surface, whereas mAb 3D11 detected HLA-F on the cell surface. The presence of HLA-F on T cells was recognized by mAb FG1 but not by mAb Fpep1.1. mAb 3D11 detected HLA-F on the cell surface of activated B cells and on peripheral blood lymphocytes, but not on the normal cells. Importantly, mAb 3D11 revealed that HLA-F exists as a heavy chain (HC) monomer, rather than as an HC associated with B2m. Although these mAbs are believed to be specific to HLA-F, their monospecificity has not been formally established, which is critical for immunodiagnostic and therapeutic purposes. Previously, we investigated the diversity of HLA class I reactivities of anti-HLA-E mAbs using HLA-I coated multiplex bead assays on a Luminex platform. We reported that more than 80% of the HLA-E mAbs were cross-reactive with other HLA-I molecules, with exceptionally few truly HLA-E-monospecific mAbs. In the present investigation, we generated IgG mAbs against HCs of HLA-F in Balb/C mice and examined the cross-reactivity of anti-HLA-F mAbs with other HLA-I alleles using a multiplex bead assay on the Luminex platform. Beads coated with an array of HLA homo- and heterodimers of different HLA-Ia (HLA-A, HLA-B, and HLA-C) and Ib (HLA-E, HLA-F, and HLA-G) alleles were used to examine the binding of the anti-HLA-F mAbs. Only two mAbs were HLA-F monospecific, and five were HLA-Ib restricted. Several anti-HLA-F mAbs cross-reacted with HLA-E (n = 4), HLA-G (n = 3), HLA-Ia alleles (n = 9), HLA-G and HLA-Ia (n = 2), and HLA-Ib and HLA-Ia (n = 6). This monospecificity and polyreactivity were corroborated by the presence of HLA-F monospecific and HLA-I-shared sequences. This study emphasizes the need to monitor the mono-specificity of HLA-F for reliable immunodiagnostics and passive immunotherapy.

Keywords: B2microglobulin; HLA-E; HLA-F; HLA-G; HLA-Ia; HLA-Ib; IgG isotypes; Luminex; heavy chain; heterodimers; mean fluorescent intensity (MFI); monoclonal IgG antibodies; monomeric; monomers; monospecific; multiplex bead assays; polyreactive.

Grants and funding

This research received funding from Mark Terasaki, the first son of the late Paul Ichiro Terasaki of Terasaki Foundation Laboratory (TFL), enabled us to initiate this project and was utilized for the completion of this work.