Knockdown of the long isoform of the prolactin receptor selectively targets pathogenic immune cells in systemic lupus erythematosus and averts glomerular pathology

Abstract Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by chronic inflammation in multiple organ systems. While a clinical association between elevated levels of the hormone/cytokine, prolactin (PRL), and exacerbation of SLE has been recognized for some time, little is known about the mechanisms through which PRL affects the course of this disease. Here, we show that immune cells in SLE have aberrant splicing of the prolactin receptor (PRLR) such that the ratio of the long to short splice variants is increased. To determine whether the change in PRLR isoform expression was causal in this disease, we used a splice-modulating oligomer (SMO) that knocked down expression of the long splice variant (LFPRLR). Using patient samples ex vivo, SLE-prone mice in vivo, high-dimensional flow cytometry and single-cell RNA-sequencing, we demonstrate that the aberrant PRLR isoform expression in SLE both directly and indirectly drives production of autoreactive immune cell phenotypes. Thus, LFPRLR knockdown decreased the expression of type I interferon signaling/response genes known to be biomarkers and hub genes for SLE, reduced immunoglobulins with signatures considered autoreactive, and averted glomerular kidney damage in SLE-prone mice. Importantly, LFPRLR knockdown reduced pathogenic B cells and other immune subsets that drive B-cell activation, without negative impact on healthy donor counterparts. Current treatments for SLE adversely affect healthy cells and do not concurrently eradicate multiple pathogenic immune subsets. Since LFPRLR SMO does not share these disadvantages, knockdown of the LFPRLR represents a potential treatment strategy for SLE that merits further investigation. Graphical Abstract The LFPRLR represents an attractive therapeutic target in SLE. (Left) In addition to increased pituitary/circulating PRL, individuals with SLE exhibit aberrant increases in the production of autocrine/paracrine PRL by immune cells, and in their expression of specifically the long isoform (LF) of the PRLR. (Middle) Expression of the LFPRLR specifically enhances autoreactive immunophenotypes and promotes lupus nephritis. (Right) A splice modulating oligomer (SMO), that prevents synthesis of only the LFPRLR but not the short PRLR isoforms, reduces pathogenic immunophenotypes without affecting the normal counterparts of immune cells. Competing Interest Statement The authors have declared no competing interest. Abbreviations - aa - amino acid - APRIL - A proliferation-inducing ligand - BAFF - B-cell activating factor - BCR - B-cell receptor - CD - cluster of differentiation - CDR3 - complementary determining region 3 - CM - classical monocytes - DAPI - 4′,6-diamidino-2-phenylindole - DC - dendritic cell - DMSO - dimethyl sulfoxide - DNA - deoxyribonucleic acid - DSMZ - Deutsche Sammlung von Mikroorganismen und Zellkulturen - EdU - 5-ethynyl-2′-deoxyuridine - FAS - fas cell surface death receptor - FBS - fetal bovine serum - FPKM - fragments per kilobase of transcript per million mapped reads - GC - germinal center - GSEA - gene set enrichment analysis - IACUC - Institutional Animal Care and Use Committee - IC50 - half-maximal inhibitory concentration - IFN-I - type I interferon - IFPRLR - intermediate isoform of PRLR - IGH - immunoglobulin heavy chain - JAK - Janus kinase - JAX - Jackson Laboratories - LFPRLR - long isoform of PRLR - MACS - magnetic activated cell sorting - MFI - median fluorescence intensity - mRNA - messenger ribonucleic acid - MRL/lpr - Murphy Roths Large lymphoproliferation - MTS - (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) - I/NCM - intermediate and non-classical monocytes - NGS - next generation sequencing - NK - natural killer - PBMC - peripheral blood mononuclear cells - PRL - prolactin - PRLR - prolactin receptor - RNA-seq - RNA sequencing - scRNA-seq - single cell RNA sequencing - SFPRLR - short isoform of PRLR - SLE - systemic lupus erythematosus - SMO - splice modulating oligomer - STAT - signal transducer and activator of transcription - UB - ubiquitin - WBC - white blood cell - UMI - unique molecular identifiers - NES - normalized enrichment score - siRNA - small interfering RNA.