lncRNA and Multiple Sclerosis

Matteo Lucchini

PhD student and neurologist at Fondazione Policlinico

Universitario Agostino Gemelli IRCCS, Università Cattolica

del Sacro Cuore - Roma

• Most common cause of neurological disability among young people

• Women>men (3:1 ratio)

• Typical onset between 20 to 40 years

• Approximately 2 million people are diagnosed with MS globally

• Estimated median incidence is 2.5 per 100,000

• The incidence of MS varies geographically (latitudinal gradient)

• Wide clinical variability

• 85% relapsing-remitting course at onset (nearly an half eventually developing progressive course within 20 years)

• 15% primary progressive course

Multiple Sclerosis

Multiple Sclerosis

Several treatment options with different immunological targets

Several known risk factors: genetic, ambiental and behaviour susceptibility such as low vitamin D levels, EBV infections, female sex, smoke, obesity and risk genes

Multiple Sclerosis

Evolving diagnostic criteria (McDonald 2017 revision) allowing early diagnosis based on clinical and radiological findings

In the last revision new relevance to CSF-restricted Oligoclonal bands (OCBs)

Multiple Sclerosis

Complex physiopathology primarlyinvolving adaptative immune system

Interplay between autoreactive T cells (Th17 and Th1), B cells with macrophagic and microglialactivation

Inbalance between pro-inflammatoryand anti-inflammatory pathways

Currently available treatment options exert different and specific effects on immune system

- Immunomodulating agents

- Sequestrant agents

- Depletive agents

Multiple Sclerosis and sncRNAs

• 650 dysregulated miRNA (at least one study) 179 at least two studies 58 at least three studies 8 at least four studies

• Heterogeneity between samples, cohorts and methodologies

• TGF-b signaling is the most significant pathway affected (involved in proliferation of Th1 and Th17 cells and Treg differentiation, upregulated miR-142, miR-145 and miR-210)

• Upregulated miR-155 in T cells and PBMCs: promoted development and differentiation of Th1 and Th17 cells; reduced after specific treatments

• Upregulated miR-326 in EAE: promotion of Th17 differentiation

• Targeting miR-155 and miR-326 in EAE model produce a milder phenotype

• Circulating miR-150 and miR-181c as possible disease biomarkers

• miR-320, miR-320b and miR-629 for PML risk profile

Multiple Sclerosis and lncRNA

• 28 papers from 2016 explored the relationship between specific lncRNA subsets and MS

• Different sources explored: serum, PBMCs, microglial and biopsy/autopsy samples from MS patients; EAE mouse model

• Array analysis

• Expression of putative lncRNAs involved in specific immunological processes

• Association between MS and specific lncRNA polymorphisms (only one cohort)

• Cohorts including non treated and treated (exclusively IFNb-1a) MS patients

• Mainly RRMS patients; only few papers includes some progressive MS

• 11 papers from the same cohort

- TUG1 may be a component of p53-regulatory network participating to the apoptotic pathways active in early stages of neurodegenerative and inflammatory diseases

- NEAT1 is induced by activation of TLR3-p38 pathway; high levels of NEAT1 promote the transcriptional activation of IL-8, that is significantly high in MS patients

- RN7SK RNA was part of the 7SK snRNP complex that can repress the cellular kinase complex P-TEFb, involved in CD4+ T lymphocytes activation

- RN7SK RNA was up-regulated also in idiopathic inflammatory myopathy patients- TUG1 and NEAT1 upregulated in whole blood samples from an Iranian cohort

(Dastmalchi 2018, Multiple Sclerosis and related disorders)

Multiple Sclerosis and lncRNA

- PCR array with 90 common lncRNA in PBMCs from MS patients- 2 lncRNAs downregulated validated in independent cohort: NRON and

TUG1

Multiple Sclerosis and GAS5

• Growth arrest specific 5 (GAS5) accumulates in cells during growth arrest modulating apoptosis and interferes with glucorticoid receptor (GR) and glucorticoid response element (GRE) regulation

• GAS5 upregulated in blood samples of RRMS and correlated with NR3C1 expression (its nuclear target gene)

• GAS5 rs2067079 SNP, located in the promoter region, is more frequent in MS patients

Multiple Sclerosis and GAS5

• GAS5 upregulation in microglia suppresses microglial M2 polarization through TRF4 inhibition (independently from GR and GRE)

• Explored in EAE model and microglial cells from MS patients

• GAS5 interference promotes remyelination following lysolecithin-induceddemyelination

Multiple Sclerosis and MALAT1

• Metastasis associated lung adenocarcinoma transcript 1 (MALAT1) regulates leukocyte differentiation and function and is highly expressed in CNS tissues

• MALAT1 downregulated in MS brain and spinal cord EAE samples (Masoumi 2019, Journal of Neuroimmunology)

• MALAT1 also downregulated in macrophages and T cells, influencing the balance between Th1/Th17 and Tregs (Masoumi 2019, Journal of Neuroimmunology)

• Silencing MALAT1 promotes Th1/Th17 differentiation and reduces Tregs (Masoumi2019, Journal of Neuroimmunology)

• MALAT1 SNPs and MS risk: G allele rs619586 SNP less frequent in MS; A allele rs619586 and T allele rs3200401 associated with MS (Eftekharian 2019, Immunological investigations)

• MALAT1 also modulates endogenous expression of slicing factors and alternative splicing of MS-associated genes (such as IL7R and SP140) (Cardamone 2019, Human Molecular Genetics)

• MALAT1 is found upregulated in serum of MS patients and strongly correlates with lnc-DC (Shaker 2019, Bioscience reports)

Multiple Sclerosis and HOTAIR

• HOX transcript antisense intergenic RNA (HOTAIR) higher expression in whole blood sample of MS patients, reduced after vitamin D supplementation

• HOTAIR decreased in spinal cord and cortex and increased in cerebellum of EAE mice

(Kakhki 2017, Immunology)

HOTAIR modulation in EAE mouse model under different condition: sulfasalazine as promyelinating agent and cuprizone as demyelination inducer

• HOTAIR modulates a M1 pro-inflammatory polarization of microglia through interaction with miR-136-5p and NF-kB pathway activation

(Duan 2018, Biochemical pharmacology)

Multiple Sclerosis and linc-MAF4

• linc-MAF4 upregulated in PBMCs from MS compared to controls and strongly correlates with MAF downregulation

• Th1-like stimulation promotes linc-MAF4 expression

• Th2-like stimulation promotes MAF expression

• linc-MAF4 overexpression leads to Th1 differentiation and its downregulation guides to a Th2 phenotype

• linc-MAF4 correlates with annual relapse rate and MRI lesion load

(Zhang 2017, FASEB Journal)

Multiple Sclerosis and IFNg/TNFa

• GSTT1-AS1 and IFNG-AS1 inhibit IFNg and TNFa gene expression

• Those lncRNAs were found downregulated in whole blood sample of RRMS patients (IFNg and TNFa were upregulated in the same cohort)

• Expression GSTT1-AS1 / IFNG-AS1 and IFNg / TNFa genes were correlated

(Ganji 2019, Neurol Sci)

Aim: explores Th1 related lncRNAs expression in MS

• IFNG-AS1-001 and AC007278.2 upregulated in PMBCs from relapsing MS patients

• IFNG-AS1-002 upregulated in PMBCs from remitting MS patients

(Hosseini 2019, J Cell Physiol)

lncRNAs as potential biomarkers for MS?

- HULC downregulated (whole blood sample)- Role in immune system not clarify- Significative differences regarding age and gender

- HUR1 and OIP5-AS1 upregulated in whole blood- Suppose to be involved in Th17 celldifferentiation

Progressive Multiple Sclerosis and lncRNA

- TUG1 was up-regulated in SP-MS- down-regulation of non-protein coding RNA 188 (LRRC75A-AS1) and a significant up-

regulation of long intergenic non-protein coding RNA 293 (LINC00293) and RP11-29G8.3 in PP-MS

unpublished data

Conclusion

• Multiple Sclerosis has a complex physiopathology involving both adaptative and innate immune systems with a complex interplay within reparative and degenerative mechanism in CNS

• Only few studies regarding lncRNAs exploring different samples (serum, whole blood, PBMCs, EAE model, biopsy/autopsy)

• Differences regarding patients’ cohort: treated vs untreated; stable vs relapsing; only few reports regarding progressive patients

• lncRNAs dysregulated are involved in lymphocyte or microglial activation and differentiation (GAS5, MALAT1, HOTAIR, linc-MAF4)

Future perspective:

• Explore and confirm potential role of specific lncRNAs in MS pathology

• Evaluate the modulation of such lncRNAs under treatment and explore their potential as markers of response

GRAZIE PER L’ATTENZIONE!

Centro SM PoliclinicoGemelli:

Prof. M. Mirabella

Dr.ssa V. Nociti

Dr.ssa C. De Fino

Dr. F.A. Losavio

Dr.ssa M. Loiodice

Fondazione Don Carlo Gnocchi:

Dr. M. Santoro