Antigen-specific B cell depletion for precision therapy of mucosal pemphigus vulgaris
Jinmin Lee, … , Michael C. Milone, Aimee S. Payne
Jinmin Lee, … , Michael C. Milone, Aimee S. Payne
Published August 20, 2020
Citation Information: J Clin Invest. 2020;130(12):6317-6324. https://doi.org/10.1172/JCI138416.
View: Text | PDF
Concise Communication Autoimmunity

Antigen-specific B cell depletion for precision therapy of mucosal pemphigus vulgaris

Abstract

Desmoglein 3 chimeric autoantibody receptor T cells (DSG3-CAART) expressing the pemphigus vulgaris (PV) autoantigen DSG3 fused to CD137-CD3ζ signaling domains, represent a precision cellular immunotherapy approach for antigen-specific B cell depletion. Here, we present definitive preclinical studies enabling a first-in-human trial of DSG3-CAART for mucosal PV. DSG3-CAART specifically lysed human anti-DSG3 B cells from PV patients and demonstrated activity consistent with a threshold dose in vivo, resulting in decreased target cell burden, decreased serum and tissue-bound autoantibodies, and increased DSG3-CAART engraftment. In a PV active immune model with physiologic anti-DSG3 IgG levels, DSG3-CAART inhibited antibody responses against pathogenic DSG3 epitopes and autoantibody binding to epithelial tissues, leading to clinical and histologic resolution of blisters. DSG3 autoantibodies stimulated DSG3-CAART IFN-γ secretion and homotypic clustering, consistent with an activated phenotype. Toxicology screens using primary human cells and high-throughput membrane proteome arrays did not identify off-target cytotoxic interactions. These preclinical data guided the trial design for DSG3-CAART and may help inform CAART preclinical development for other antibody-mediated diseases.

Authors

Jinmin Lee, Daniel K. Lundgren, Xuming Mao, Silvio Manfredo-Vieira, Selene Nunez-Cruz, Erik F. Williams, Charles-Antoine Assenmacher, Enrico Radaelli, Sangwook Oh, Baomei Wang, Christoph T. Ellebrecht, Joseph A. Fraietta, Michael C. Milone, Aimee S. Payne

×

Figure 4

High-throughput membrane proteome array (MPA) plus targeted cell screening did not identify off-target DSG3-CAART cytotoxic interactions.

Options: View larger image (or click on image) Download as PowerPoint
High-throughput membrane proteome array (MPA) plus targeted cell screeni...
(A) More than 5300 membrane proteins were expressed in HEK293T cells, and then screened with soluble Fc-tagged proteins to assess binding. Soluble Fc-tagged DSG3-CAAR ectodomain bound weakly to CLEC4M. Px44, anti-DSG3 positive control; FCGR1A, anti-Fc internal control. (B) MPA validation, indicating DSG3-CAAR-Fc binding to CLEC4M-overexpressing HEK293T relative to Fc-isotype control. (C) K562 cells were transduced with lentivectors encoding CLEC4M isoform v1 or v8, sorted to select for high CLEC4M expression, and then incubated with DSG3-CAART or CART19 cells. IFN-γ levels were undetectable in culture supernatants after 16 hours of coincubation of DSG3-CAART and CLEC4M-K562 cells, whereas IFN-γ was detected in positive control cocultures of DSG3-CAART with Nalm6 B cells expressing anti-DSG3 BCR PVB28, or CART19 cells cocultured with CD19+ Nalm6 cells. (D) qPCR verifies CLEC4M mRNA expression in pulmonary microvascular but not hepatic sinusoidal endothelial cells. HaCat keratinocytes, negative control. (E) Luminex cytokine analysis performed on DSG3-CAART and primary human cell coculture supernatants indicates no cytotoxic cytokine (IFN-γ, TNF-α) production. Red/blue × = below/above the range of detection. (F) xCELLigence cellular impedance assay of DSG3-CAART and primary human cell cocultures indicates no detectable cytolysis of CLEC4M-expressing pulmonary microvascular endothelial cells. DSG3-CAART induced changes in hepatic endothelial cell impedance only at a 50:1 effector-to-target ratio, a ratio not likely to be achieved in vivo.