WO2024210835A1 - Method of treating psoriasis - Google Patents

Abstract

The present invention relates to the use of a therapeutically effective amount of microRNA-101-3p (miR-101-3p) for treating a subject suffering from psoriasis via the modulation of expression of Enhancer of Zeste homolog 2 (EZH2).

Description

METHOD OF TREATING PSORIASIS

RELATED APPLICATIONS

This application claims the benefit of priority of Singapore provisional application no. 10202300938Q, filed on 05 April 2023, the contents of it being hereby incorporated by reference in its entirety for all puiposes.

TECHNICAL FIELD

The present disclosure relates to the field of biotechnology. In particular, the present invention refers to non-coding nucleic acids capable of modulating gene expression. More particularly, the present disclosure relates to the modulation of Enhancer of Zeste homolog 2 (EZH2) by microRNA 101 (miR-101).

BACKGROUND

[001] Epidermal hyperplasia, a characteristic feature of psoriatic skin lesions, is epigenetically driven by Enhancer of Zeste homolog 2 (EZH2). EZH2 and EZH2-mediated trimethylation of histone H3 lysine 27 (H3K27me3) are both abnormally upregulated in psoriatic lesions.

[002] Psoriatic patients are treated symptomatically only with topical or intravenous steroids that are unable to provide long term relief, where patients have frequent flare-ups. There is thus an unmet need for managing psoriasis effectively.

SUMMARY

[003] In one aspect, the present disclosure provides a method of treating psoriasis, wherein the method comprises administering a therapeutically effective amount of a microRNA (miRNA) comprising a sequence as set forth in SEQ ID NO: 2 to a subject suffering from psoriasis.

[004] In one example, the miRNA comprises a sequence as set forth in SEQ ID NO: 3. In another example, the miRNA comprises a sequence as set forth in SEQ ID NO: 4. In another example, the miRNA comprises a sequence as set forth in SEQ ID NO: 5.

[005] In another example, the therapeutically effective amount of miRNA is a mimic of a wild-type miR-101-3p.

[006] In another example, administration of the miR-101-3p mimic decreases EZH2 expression in the subject. In yet another example, the decrease in EZH2 expression decreases psoriatic lesions in the subject suffering from psoriasis.

[007] In another aspect, the present disclosure provides a miR-101-3p mimic, wherein the miR-101- 3p mimic does not comprise a sequence as set forth in SEQ ID NO: 5.

BRIEF DESCRIPTION OF DRAWINGS

[008] miR-101-3p is significantly downregulated in psoriasis [009] Fig. 1A is a heatmap showing miRNA microarray data from biopsies of healthy skin and lesional skin from psoriasis patients. Hierarchical clustering shows the distinct expression patterns of microRNAs in lesional skin compared to healthy skin. The heatmap shows that the number of upregulated miRNAs and the number of downregulated miRNAs in lesional skin compared to healthy skin are 42 and 26 respectively.

[0010] Fig. IB is a subset heatmap showing the top 10 upregulated miRNAs and top 10 downregulated miRNAs in lesional skin compared to healthy skin. Fig. 1 B shows that microRNA 101 -3p (miR-101 -3p) is downregulated in lesional skin.

[0011] Fig. 1C is a qRT-PCR validation showing the transcript abundance of microRNA 199b (miR-199b) in total RNA isolated from lesional skin (n=5) and from healthy skin (n=5) from the patient cohort.

[0012] Fig. ID is a qRT-PCR validation showing the transcript abundance of microRNA 21 (miR-21) in total RNA isolated from lesional skin (n=5) and from healthy skin (n=5) from the patient cohort.

[0013] Fig. IE is a qRT-PCR validation showing the transcript abundance of miR-101-3p in total RNA isolated from lesional skin (n=5) and from healthy skin (n=5) from the patient cohort. Fig. IE shows that the transcript abundance of miR-101 -3p is high in healthy skin and low in lesional skin.

[0014] Fig. IF is an in situ hybridization for mature miR-101-3p in healthy and lesional skin sections (top panel, n=8). Healthy and lesional skin sections were co-stained for keratin 10 in the bottom panel. A magnified view of the indicated inset is shown in the middle column. DAPI was used as a counterstain. Fig. IF shows that miR-101-3p is found in lower levels in lesional skin compared to healthy skin. Scale bar is 20 pm.

[0015] IL-17 mediated downregulation of miR-101-3p upregulates EZH2 and promotes epidermal hyperplasia.

[0016] Fig. 2A shows the binding site for miR-101-3p on the wild-type (WT) sequence of EZH2 3’-UTR (boxed up). The seed sequence of EZH2 (boxed up) was mutated to disrupt the binding site for miR-101 -3p, where miR-101-3p binds to EZH2 via the boxed up portion.

[0017] Fig. 2B is a bar graph showing normalized luciferase activity for N/TERT-1 cells co-transfected with luciferase reporter constructs containing wild-type or mutant EZH2 3’-UTRs in the presence of miR-101-3p mimic or control. Renilla luciferase was used as a normalization control. Luciferase activity is expressed as the mean relative to the controls (n = 3). **P < .001. The student t test was used to calculate P value; error hars denote means ± SEMs. Fig. 2B shows that luciferase reporter constructs containing wild-type EZH2 3’-UTR had significantly reduced luciferase reporter activity. In contrast, luciferase reporter constructs containing mutant EZH2 3’-UTR where the miR-101-3p mimic binding site was mutated showed no change in reporter activity, demonstrating the binding region of miR-101-3p mimic to EZH2.

[0018] Fig. 2C is a confocal image showing in situ hybridization for mature miR-101 -3p and immunohistochemical staining for EZH2 in biopsies of lesional skin from psoriasis patients and healthy skin samples. DAPI (blue) is a nuclear counterstain. Fig. 2C shows that miR-101-3p is found in decreased amounts in lesional skin compared to healthy skin. Conversely, EZH2 is found in high amounts in lesional skin compared to healthy skin. Scale bar is 20 pm. [0019] Fig. 2D is a western blot showing the differences in protein expression levels of EZH2 in total cell extracts from N/TERT-1 cells transfected with control mimic, miR-101-3p mimic, or miR-144 mimic. GAPDH was used as a loading control. Fig. 2D is a representative western blot of three independent experiments. Fig. 2D shows that the protein expression level of EZH2 is lower in N/TERT-1 cells transfected with miR-101-3p mimic compared to control mimic or miR-144 mimic.

[0020] Fig. 2E shows the relative transcript abundance of miR-101 -3p in confluent N/TERT-1 cultures treated with IFNy’ 1L-17A/F, or TNFa for 24 h as determined by qRT-PCR. Fig. 2E shows that the relative transcript abundance of miR-101-3p is lowest in N/TERT-1 cells treated with IL-17A/F compared to N/TERT-1 cells treated with control, IFN\ or TNFa.

[0021] Fig. 2F is a western blot showing the levels of EZH2 expression in N/TERT-1 cultures treated with IFNyj IL-17A/F or TNFa. Fig. 2F shows that the levels of EZH2 expression is highest in N/TERT- 1 cultures treated with IL-17A/F compared to control, IFN\ . or TNFa.

[0022] Fig. 2G is a graphical model depicting the regulatory pathway of IL- 17. In normal skin, miR- 101-3p causes de-repression of EZH2 which promotes epidermal differentiation and homeostasis. In lesional skin, increased IL-17 activity downregulates miR-101-3p which is necessary for de-repression of EZH2. Increased amounts of EZH2 increases EZH2-mediated H3K27 trimethylation which leads to keratinocyte proliferation and epidermal hyperplasia.

DETAILED DESCRIPTION

[0023] Psoriasis is a chronic, systemic autoimmune disease. Persistent inflammation, uncontrolled keratinocyte proliferation and aberrant differentiation are the major hallmarks of psoriasis. Subjects suffering from psoriasis commonly display symptoms of lesions. Lesions can be discrete, oval erythematous papules or plaques covered with thick, silvery scales. Lesions can also be accompanied by itching and are localized on external skin, such as the knees, elbows, trunk, or scalp. While psoriasis is rarely life-threatening, the lesions resulting from psoriasis can be detrimental to a subject’s quality of life. [0024] Excessive keratinocyte proliferation, coupled with poor differentiation, is a characteristic feature of psoriatic skin lesions. This hyperproliferative phenotype is caused by epigenetic factors - particularly EZH2 (Enhancer of Zeste homolog 2) and EZH2 -mediated trimethylation of lysine 27 on histone H3 (H3K27me3). Both EZH2 and H3K27me3 which are overexpressed in carcinoma are also upregulated in psoriatic lesions.

[0025] Current treatments of psoriasis are focused on managing symptoms by means of topical steroids but arc unable to provide long-term relief. There is thus an unmet need for a therapeutic treatment of psoriasis.

[0026] Epigenetic pathways are known to be linked to the regulation of gene expression, where regulatory mechanisms are mediated by non-coding RNAs. As used herein, the term “non-coding RNA” refers to RNA molecules that are not translated into a protein but can have functional roles in transcription and translation. Examples of non-coding RNAs can be, but are not limited to, microRNAs (miRNAs), small interfering RNAs (siRNAs), piwi-interacting RNAs (piRNAs), transfer RNAs (tRNAs), or ribosomal RNAs (rRNAs). In one example, the non-coding RNA is a miRNA.

[0027] As used herein, the term “miRNA” refers to small, single-stranded, non-coding RNA molecules containing 21 to 23 nucleotides and are involved in RNA silencing and post-transcriptional regulation of gene expression, where they can suppress translation of their target transcripts. Examples of miRNA can he, but are not limited to, miR-101 , miR-199, miR-31 , miR-125b, miR-21 , or miR-203. In one example, the miRNA is miR-101. In another example, the miRNA is miR-101-3p.

[0028] Many diseases are linked to dysfunctional gene expression. In atopic dermatitis, keratinocyte differentiation is epigenetic ally regulated by miRNA.

[0029] As described herein, analysis of differentially expressed miRNAs in lesional skin from psoriasis patients identified miR-101 to be a miRNA that was consistently downregulated in psoriatic lesions compared to healthy skin. An inverse correlation between the expression of miR-101 and EZH2 was observed in psoriatic lesional skin.

[0030] Accordingly, in one aspect, the disclosure provides herein a method of treating psoriasis, wherein the method comprises administering a therapeutically effective amount of a microRNA (miRNA) comprising a sequence as set forth in SEQ ID NO: 2 to a subject suffering from psoriasis. In one example, the miRNA comprises a sequence as set forth in SEQ ID NO: 3. In another example, the miRNA comprises a sequence as set forth in SEQ ID NO: 4. In yet another example, the miRNA comprises a sequence as set forth in SEQ ID NO: 5.

[0031] In another aspect, provided herein is a use of a miRNA comprising a sequence as set forth in SEQ ID NO: 2 in the manufacture of a medicament for treating psoriasis. In one example, the miRNA comprises a sequence as set forth in SEQ ID NO: 3. Tn another example, the miRNA comprises a sequence as set forth in SEQ ID NO: 4. In yet another example, the miRNA comprises a sequence as set forth in SEQ ID NO: 5.

[0032] In another aspect, provided herein is a miRNA comprising a sequence as set forth in SEQ ID NO: 2 for use in treating psoriasis. In one example, the miRNA comprises a sequence as set forth in SEQ ID NO: 3. In another example, the miRNA comprises a sequence as set forth in SEQ ID NO: 4. In yet another example, the miRNA comprises a sequence as set forth in SEQ ID NO: 5.

[0033] In yet another aspect, provided herein is a miRNA comprising a sequence as set forth in SEQ ID NO: 2 for use in therapy. In another aspect, provided herein is a miRNA comprising a sequence as set forth in SEQ ID NO: 2 for use in medicine. In one example, the miRNA comprises a sequence as set forth in SEQ ID NO: 3. In another example, the miRNA comprises a sequence as set forth in SEQ ID NO: 4. In yet another example, the miRNA comprises a sequence as set forth in SEQ ID NO: 5.

[0034] To ensure that the miRNA administered to a subject is able to decrease EZH2 expression, the miRNA should be capable of binding to the target protein EZH2. Thus, in one example, the miRNA as used herein is a mimic of a wild-type miR-101. In another example, the miR-101 mimic as used herein is a mimic of miR-101-3p. As used herein, the term “mimic” refers to a synthetic RNA construct which has substantially the same sequence as an endogenous RNA and performs substantially the same function.

[0035] As used herein, the term ‘’substantially the same sequence” in the context of the miR-101 mimic as used herein refers to a mimic which has a nucleotide sequence that is similar to or identical to the wildtype miR-101. In one example, the miR-101 mimic is a mimic of wild-type miR-101-3p. In another example, the miR-101 mimic has one or more nucleotide differences with the sequence of wild-type miR-101-3p. In another example, the sequence of the miR-101-3p mimic has no nucleotide differences with the sequence of wild-type miR-101-3p. In yet another example, the miR-101-3p mimic is identical to the wild-type miR-101-3p.

[0036] Accordingly, in one example, the miRNA as described herein comprises a sequence as set forth in SEQ ID NO: 2, where the miRNA has 13 nucleotide differences or less compared to the sequence of wild-type miR-101-3p. In another example, the miRNA as described herein comprises a sequence as set forth in SEQ ID NO: 3, where the miRNA has 9 nucleotide differences or less compared to the sequence of wild-type miR-101-3p. In another example, the miRNA as described herein comprises a sequence as set forth in SEQ ID NO: 4, where the miRNA has 6 nucleotide differences or less compared to the sequence of wild-type miR-101-3p. In another example, the miRNA as described herein comprises a sequence as set forth in SEQ ID NO: 5, where the miRNA has no nucleotide differences compared to the sequence of wild-type miR-101-3p.

[0037] Thus, in one example, the miR-101 mimic has a sequence that has substantially the same sequence as wild-type miR-101-3p. In a further example, the miR-101-3p mimic is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95%, identical to SEQ ID NO: 1.

[0038] In another example, the miR-101 -3p mimic does not comprise a sequence as set forth in SEQ ID NO: 5.

[0039] As used herein, the term “nucleotide difference” refers to a modification of a nucleotide sequence compared to a reference sequence, where examples of modification can be, but are not limited to, base substitutions, base deletions, base additions, base methylation, base acetylation, locked nucleic acids (LNAs), phosphorothioate backbone modifications, and any other modifications known to a skilled person.

[0040] Thus, in one example, the miR-101-3p mimic comprises a sequence as set forth in SEQ ID NO: 2, where the miR-101-3p mimic as described herein has 13 nucleotide differences or less compared to the sequence of wild-type miR-101 -3p. In another example, the miR-101 -3p mimic as described herein comprises a sequence as set forth in SEQ ID NO: 3, where the miR-101-3p mimic has 9 nucleotide differences or less compared to the sequence of wild-type miR-101-3p. In another example, the miR- 101 -3p mimic as described herein comprises a sequence as set forth in SEQ ID NO: 4, where the miR- 101-3p mimic has 6 nucleotide differences or less compared to the sequence of wild-type miR-101-3p. In another example, the miR-101-3p mimic as described herein comprises a sequence as set forth in SEQ ID NO: 5, where the miR-101-3p mimic has no nucleotide differences compared to the sequence of wild-type miR-101-3p.

[0041] It is understood by a skilled person that a miR-101-3p mimic can bind to EZH2, and it is also understood by a skilled person that a miR-101-3p mimic has substantially the same sequence as an endogenous wild-type miR-101-3p, where the similarity in sequence between a miR-101-3p mimic and a wild-type miR-101 -3p can be located on a binding region. As used herein, the term “binding region” refers to a region on a first nucleotide sequence which binds to a region on a second nucleotide sequence by complementary base pairing. It is understood by a skilled person that the binding of a first nucleotide sequence to a second nucleotide sequence at such regions are sufficient for binding of the entirety of the first nucleotide sequence to a second nucleotide sequence, even if other regions on the first and second nucleotide sequence are not complementary to each other. Accordingly, the miR-101-3p mimic as used herein binds to EZH2 via the binding region. In one example, the sequence as set forth in SEQ ID NO: 6 is the binding region of miR-101-3p. In another example, the binding region is called a seed sequence.

[0042] To ensure successful delivery of the miRNA as described herein to a subject, the miRNA should be formulated and administered suitably.

[0043] In one example, the miRNA as described herein can be packaged with or within a carrier selected from a pharmaceutically acceptable excipient, a transfection agent, a liposome, a conjugate, a nanoparticle, an exosome, a saline solution, or any other suitable carrier which will be known to a skilled person.

[0044] In another example, to formulate the miRNA suitably for topical administration, the miRNA formulation can further comprise preservatives, buffers, stabilizing agents, or any other suitable additive for formulating the miRNA to be suitable for topical administration. Examples of suitable additives can be, but are not limited to, animal and vegetable fats, oils, waxes, paraffins, emulsifiers, emollients, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc and zinc oxide, or any other suitable additive known to a skilled person for formulating a topical composition. In a further example, the miRNA is formulated as a cream, an ointment, a spray, a paste, a powder, a foam, a lotion, a gel, a solution, a patch, a composition, a formulation, a suspension, a solution, an emulsion, or any other forms suitable for topical administration which are known in the art.

[0045] In yet another example, to formulate the miRNA suitably for systemic administration, the miRNA formulation can further comprise additives or diluents such as a binder, a solubilizing agent, a suspension aid, an emulsifying agent, a stabilizing agent, a pH adjusting acid, base, or buffers, or any other suitable additive or diluents for formulating the miRNA to be suitable for systemic administration. In a further example, the miRNA is formulated as an infusion, an injection, a composition, a formulation, a suspension, a solution, or any other forms suitable for systemic administration which are known in the art. [0046] In another example, the miRNA as described herein is provided as a kit. Such kits can further comprise carriers, additives, or instructions for use of the miRNA as described herein.

[0047] The amount of miRNA to be administered can be calculated by a skilled person. The calculation of the amount of miRNA to be administered can be performed with factors such as, but are not limited to, age, weight, severity of psoriatic lesions and disease, the mode of administration, and any other factors known by a skilled person to be important for calculating the amount of miRNA to be administered to increase the level of miRNA in a subject. In one example, the amount of miRNA to be administered is between 0.1 to 1 pM/kg of body weight, 1 to 100 pm/kg of body weight, 100 pm to 1 mM/kg of body weight, 1 mM to 10 mM/kg of body weight, 10 mM to 100 mM/kg of body weight, or 100 mM to 1 M/kg of body weight.

[0048] Methods of administration are well-known in the art. In one example, the miRNA can be administered systemically. Examples of systemic administration can be, but are not limited to, intravenous, intraperitoneal, intramuscular, subcutaneous, or any other mode of systemic administration known in the art.

[0049] In one example, administration of the miRNA as described herein results in an increase in the level of miR-101-3p in a subject, where the increase in the level of miR-101-3p is sufficient to decrease expression of EZH2 in the subject. Thus, in another example, administration of the therapeutically effective amount of miRNA as described herein decreases EZH2 expression in a subject. As used herein, the term “decreased EZH2 expression” in the context of miR-101 -3p administration to a subject refers to a reduction in the level of EZH2 protein or mRNA in a subject compared to before the subject was administered miR-101-3p. Decreased EZH2 expression can be measured by any methods known in the art, which can be, but are not limited to, qRT-PCR, western blotting, in situ hybridization, confocal imaging, luciferase assay, heat mapping, and any other methods known in the art.

[0050] In a further example, the decrease in EZH2 expression decreases psoriatic lesions in a subject suffering from psoriasis. As used herein, the term “decreased psoriatic lesions” in the context of miR- 101-3p administration to a subject refers to a reduction in the severity of psoriatic lesions compared to before the subject was administered miR-101-3p. Methods of ascertaining a decrease in severity are known in the art, which can be, but are not limited to, a clinical examination, a biopsy, an observation of morphology of plaque type, and any other methods known in the art, where a decrease in severity is demonstrated by, for example, a reduction in the extent of lesions on the external skin surface, a decrease in itching, and any other reduction in symptoms of psoriasis that can be easily determined by a skilled person.

[0051] The disclosure as described herein may suitably be practiced in the absence of any element or elements, limitation or limitations, not specifically disclosed herein. Thus, for example, the terms "comprising", "including", "containing", etc. shall be read expansively and without limitation. Additionally, the terms and expressions employed herein have been used as terms of description and not of limitation, and there is no intention in the use of such terms and expressions of excluding any equivalents of the features shown and described or portions thereof, but it is recognized that various modifications are possible within the scope of the invention claimed. Thus, it should be understood that although the present disclosure has been described by preferred examples and optional features, modification and variation of the examples embodied therein herein disclosed may be resorted to by those skilled in the art, and that such modifications and variations are considered to be within the scope of this disclosure.

[0052] Certain examples may also be described broadly and generically herein. Each of the narrower species and sub-generic groupings falling within the generic disclosure also form part of the disclosure. This includes the generic description of the embodiments with a proviso or negative limitation removing any subject matter from the genus, regardless of whether or not the excised material is specifically recited herein.

EXAMPLES

[0053] Example 1: miR-101-3p is significantly downregulated in psoriasis

[0054] Global miRNA expression in lesional skin from psoriasis patients was profiled and compared to healthy skin to identify miRNA modulators of epigenetic pathways. Punch biopsies were obtained from lesional skin from 10 psoriasis patients. Punch biopsies from 10 healthy donors were obtained as a control. The clinical characteristics of patients and healthy controls are listed in Table 3

[0055] Skin miRNA transcriptomes were profiled for 4 psoriasis patients and 4 healthy controls using the miRCURY LNATM microRNA Array 7th Gen (Fig. 1A and IB). The Exiqon files pertaining to the lesional and healthy samples were imported into Partek Genomics Suite using the microRNA expression workflow. Once imported, the data was subjected to background subtraction (normalised to baseline) and LOESS transformation. The lesional samples were then compared to the healthy samples (Lesional vs Healthy) using ANOVA. Differentially expressed miRNAs were shortlisted using p-value with FDR <0.05 and a two-fold (+/-2) fold change cut-off. Around 68 significant differentially expressed miRNAs were obtained that passed the p-value with FDR (<0.05) and two-fold (+/-2) fold change cut-off. The number of upregulated miRNAs is 42 and the number of downregulated miRNAs is 26.

[0056] Expression of differentially expressed mature miRNAs in skin biopsies from healthy donors, and LS from psoriasis patients (n=6) were next measured using qRT-PCR. Relative transcript abundance of miRNAs shows the concordance between miRNA microarray data and qRT-PCR datasets (Fig. 1C, D and E). The relative expression of these miRNAs in healthy and lesional skin was further confirmed by in situ hybridization on skin sections (n=8). In healthy skin, high expression of miR-101-3p was observed in the suprabasal differentiating layers of the epidermis. The expression of miR-101-3p was reduced in psoriatic lesional skin (Fig. IE, IF).

[0057] Example 2: IL-17 mediated downregulation of miR-101-3p upregulates EZH2 and promotes epidermal hyperplasia. [0058] miR-101-3p was predicted to have a binding site in the 3’ UTR of the EZH2 transcript (Fig. 2A). 3 ’-UTR was cloned into a luciferase reporter construct and co-transfection of miR-101-3p mimics with EZH2 wild-type 3’-UTRs significantly reduced luciferase reporter activity. In contrast, when the miR-101-3p binding site was mutated, no change in reporter activity was observed, demonstrating that miR-101-3p binds to a region on EZH2 (shown boxed up in Fig. 2B). An inverse correlation between the expression pattern of miR-101 -3p and EZH2 was observed in normal skin sections unaffected by psoriasis (Figure 2C, upper panels). EZH2 expression in psoriasis is thus shown to be regulated by miR- 101-3p, where downregulation of miR-101-3p in psoriasis permits EZH2 upregulation. Increased expression of EZH2 and increased frequency of nuclear staining are observed in lesional skin sections (Fig. 2C), where the nuclear localization is consistent with EZH2’s role in histone modification. A significant decrease in the expression of EZH2 in N/TERT-1 cells transfected with miR-101-3p mimics, compared to the control mimics, is shown by Western blot analysis which shows that EZH2 is a direct target of miR-101-3p (Fig. 2D). EZH2 expression was not affected in cells transfected with miR-144 mimics which shows the specificity of miRNA targets (Fig. 2D).

[0059] Downregulation of miR-101-3p in the microenvironment of psoriatic lesions was shown to allow sustained overexpression of EZH2, which in turn drives H3K27 trimethylation and promotes keratinocyte proliferation. In lesional skin, abnormal keratinocyte gene expression profiles are supported by a cytokine milieu originating from both keratinocytes and infiltrating immune cells. Treatment of N/TERT-1 cells with IFN-y, TNF-a, and IL-17 which are the main components of milieu shows that IL-17 is a major regulator of miR-101-3p expression, where qRT-PCR shows a significant reduction in miR-101-3p expression in response to IL-17 treatment compared to the control, but not with TFN-y or TNF-a treatment (Fig. 2E). This demonstrates that IL-17 is an upstream regulator of miR- 101-3p Total cell extracts were also collected and subjected to Western blot analysis. A significant increase in EZH2 expression in response to IL- 17 treatment was observed compared to the control (Fig. 2F). The data demonstrates the role of IL- 17 as an upstream regulator of this epigenetic pathway [0060] Thus, it is shown herein that IL-17 mediates modulation of miR-101-3p expression, which in turn elicits sustained expression of EZH2, that leads to enhanced keratinocyte proliferation and epidermal hyperplasia in psoriasis (Fig. 2G).

[0061] Table 1: Sequences as described herein

[0062] Table 2: Figure legend of table 1

[0063] Table 3: Patient samples

Patients with Psoriasis (10) and Healthy controls (10) were enrolled from the National Skin Centre in Singapore in accordance with the Declaration of Helsinki and with approval by institutional review board. After giving written, informed consent, all subjects provided demographic information, including age, gender, ethnicity, medical history, and current medications. From each subject, 4 mm punch biopsies were collected from lesional skin or healthy skin respectively.

REFERENCES

[0064] Rendon, A. and K. Schakel, Psoriasis Pathogenesis and Treatment. Int J Mol Sci, 2019. 20(6). [0065] Zhang, T., et al., EZEE2-dependent epigenetic modulation of histone H3 lysine-27 contributes to psoriasis by promoting keratinocyte proliferation. Cell Death Dis, 2020. 11(10): p. 826. [0066] Quah, S., G. Subramanian, and P. Sampath, Repurposing Belinostat for Alleviation of Atopic

Dermatitis. Dermatol Ther (Heidelb), 2021 . 1 (3): p. 655-660.

[0067] Licw, W.C., ct al., Belinostat resolves skin barrier defects in atopic dermatitis by targeting the dysregulated miR-335:SOX6 axis. J Allergy Clin Immunol, 2020.

[0068] Peng, H., et al., microRN A- 31 /factor-inhibiting hypoxia-inducible factor 1 nexus regulates keratinocyte differentiation. Proc Natl Acad Sci U S A, 2012. 109(35): p. 14030-4.

[0069] Yang, Z., et al., STAT3/SH3PXD2A-ASl/miR-125b/STAT3 positive feedback loop affects psoriasis pathogenesis via regulating human keratinocyte proliferation. Cytokine, 2021. 144: p. 155535.

[0070] Meisgen, F., et al., MiR-21 is up-regulated in psoriasis and suppresses T cell apoptosis. Exp Dermatol, 2012. 21(4): p. 312-4.

[0071] Wang, M.J., et al., Role of an imbalanced rniRNAs axis in pathogenesis of psoriasis: novel perspectives based on review of the literature. Oncotarget, 2017. 8(3): p. 54985507.

[0072] Li, B., D. Xie, and H. Zhang, MicroRN A- 101 -3p advances cisplatin sensitivity in bladder urothelial carcinoma through targeted silencing EZH2. J Cancer, 2019. 10(12): p. 2628-2634.

Claims

1 . A method of treating psoriasis, wherein the method comprises administering a therapeutically effective amount of a microRNA (miRNA) comprising a sequence as set forth in SEQ ID NO: 2 to a subject suffering from psoriasis.

2. The method of claim 1, wherein the miRNA comprises a sequence as set forth in SEQ ID NO:

3.

3. The method of any one of claims 1 to 2, wherein the miRNA comprises a sequence as set forth in SEQ ID NO: 4.

4. The method of any one of claims 1 to 3, wherein the miRNA comprises a sequence as set forth in SEQ ID NO: 5.

5. The method of any one of claims 1 to 4, wherein the miRNA is a mimic of a wild-type microRNA- 101 -3p (miR-101-3p).

6. The method of any one of claims 1 to 5, wherein administration of the therapeutically effective amount of miRNA decreases EZH2 expression in the subject.

7. The method of any one of claims 1 to 6, wherein the decrease in EZH2 expression decreases psoriatic lesions in the subject suffering from psoriasis.

8. A miR-101-3p mimic, wherein the miR-101-3p mimic does not comprise a sequence as set forth in SEQ ID NO: 5.