TPP2 mutation associated with sterile brain inflammation mimicking MS

Abstract

Objective: To ascertain the genetic cause of a consanguineous family from Syria suffering from a sterile brain inflammation mimicking a mild nonprogressive form of MS.

Methods: We used homozygosity mapping and next-generation sequencing to detect the disease-causing gene in the affected siblings. In addition, we performed RNA and protein expression studies, enzymatic activity assays, immunohistochemistry, and targeted sequencing of further MS cases from Austria, Germany, Canada and Jordan.

Results: In this study, we describe the identification of a homozygous missense mutation (c.82T>G, p.Cys28Gly) in the tripeptidyl peptidase II (TPP2) gene in all 3 affected siblings of the family. Sequencing of all TPP2-coding exons in 826 MS cases identified one further homozygous missense variant (c.2027C>T, p.Thr676Ile) in a Jordanian MS patient. TPP2 protein expression in whole blood was reduced in the affected siblings. In contrast, TPP2 protein expression in postmortem brain tissue from MS patients without TPP2 mutations was highly upregulated.

Conclusions: The homozygous TPP2 mutation (p.Cys28Gly) is likely responsible for the inflammation phenotype in this family. TPP2 is an ubiquitously expressed serine peptidase that removes tripeptides from the N-terminal end of longer peptides. TPP2 is involved in various biological processes including the destruction of major histocompatibility complex Class I epitopes. Recessive loss-of-function mutations in TPP2 were described in patients with Evans syndrome, a rare autoimmune disease affecting the hematopoietic system. Based on the gene expression results in our MS autopsy brain samples, we further suggest that TPP2 may play a broader role in the inflammatory process in MS.

Figure 1. Mutation identification and Western blot

(A) Exome sequencing alignment data of the homozygous TPP2 mutation in II.1 and II.2 visualized with the IGV tool. (B) Pedigree of consanguineous Syrian family (MS01) affected with MS. Sanger chromatograms of heterozygous parents and homozygous siblings are depicted below their respective symbol. Arrow, position of mutation. (C) Homozygosity plot from Syrian family. Red bars, regions of homozygosity; arrow, homozygous block shared by all 3 siblings containing TPP2 and ERCC5. (D) Chromatograms of homozygous and heterozygous p. Thr676Ile TPP2 mutation carrier of confirmatory sequencing cohort. (E) Western blot of TPP2 in PBMCs from II.1, II.2, and 3 healthy control individuals. Black filled symbols = affected; ERCC5 = excision repair cross-complementation group 5; IGV = integrated genome viewer; mt/mt = homozygous for the mutation; open symbols = unaffected; PBMC = peripheral blood mononuclear cell; TPP2 = tripeptidyl peptidase II; wt/mt = heterozygous for the mutation.

Figure 2. TPP2 expressions in MS

(A) No expression in NAWM, while increased expression was seen in the PPWM with staining of some microglia nodules. The highest expression of TPP2 was seen in initial stages (Initial) of MS lesions, where most of the activated microglia expressed TPP2. TPP2 expression decreased from the lesion edge toward the lesion center. (B) Double staining confirmed that TPP2 expression was present in macrophages/microglia, expressing the phagocytosis-associated marker CD68 and MHC Class I and Class II molecules. (C) Quantification of TPP2-positive cell counts. Box plots displaying the number of microglia per mm² in regions of interest. Significantly more TPP2 expression was found in MS NAWM, inactive MS lesions, PPWM, initial lesions and in EA or LA lesions of MS patients compared with control white matter. ***p < 0·001, **p < 0·01, Wilcoxon test and Mann-Whitney U test. (D) TPP2 mRNA expression in brain lesions of MS patients and controls. Figure is derived from the data set GDS4218 deposited in the GEO Profile database. EA = early active; GEO Profile = gene expression omnibus profile; LA = late active; MHC = major histocompatibility complex; NAWM = normal-appearing white matter; PPWM = periplaque white matter; TPP2 = tripeptidyl peptidase II.