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MINISTRY OF EDUCATION AND TRAINING MINISTRY OF DEFENCE 108 INSTITUTE OF CLINICAL MEDICAL AND PHARMACEUTICAL SCIENCES RESEARCH ON CYTOKINE CHANGES AND TREATMENT EFFECTIVENESS OF GENERALIZED ERYTHRODERM[.]

MINISTRY OF EDUCATION AND TRAINING MINISTRY OF DEFENCE

108 INSTITUTE OF CLINICAL MEDICAL AND PHARMACEUTICAL SCIENCES

RESEARCH ON CYTOKINE CHANGES AND TREATMENT EFFECTIVENESS OF GENERALIZED ERYTHRODERMIC

THE THESIS WAS DONE IN: 108 INSTITUTE OF CLINICAL MEDICAL AND PHARMACEUTICAL SCIENCES

Supervisor:

1 Assoc.Professor Đặng Văn Em MD, PhD

2 Assoc Professor Lê Hữu Doanh MD, PhD

Reviewer:

1 Assoc.Professor Phạm Hoàng Khâm MD, PhD

2 Assoc.Professor Bùi Thị Vân MD, PhD

This thesis will be presented at Institute Council at: 108 Institute of Clinical Medical and Pharmaceutical Sciences

Day Month Year

The thesis can be found at:

1.National Library of Vietnam

2.Library of 108 Institute of Clinical Medical and Pharmaceutical Sciences

3.Central Institute for Medical Science Infomation and

Tecnology

RESEARCH PROBLEM

Generalized erythrodermic psoriasis (GEP) is one of the severe forms of psoriasis characterized by extensive skin lesions covering over 90% of the body's surface area, accompanied by disruptions in biochemical, water, and electrolyte balance, as well as damage to various organs It significantly impacts aesthetics, psychological well-being, and quality of life, burdening both families and society with healthcare responsibilities Recent studies have indicated a significant increase in cytokines such as IL-2, IL-6, IL-8, IL-10, IL-

12, IL-17, IL-23, etc., in the serum of psoriasis patients These cytokines are believed to play a role in maintaining psoriatic lesions

in general and GEP in particular These cytokines can be used as monitoring and treatment indicators for GEP patients in particular and psoriasis patients in general Several biologic drugs have been studied and applied clinically, including Infliximab-TNF-α inhibitor and Secukinumab - IL-17 inhibitor While numerous studies on GEP and post-treatment cytokine evaluation have been conducted worldwide, there is still a lack of research on post-treatment cytokine evaluation In Vietnam, there have been studies on cytokines in common psoriasis and pustular psoriasis, but none have explored the association of cytokines (IL-2, IL-4, IL-6, IL-8, IL-10, IL-17, TNF-α, INF-γ) in GEP patients before and after Methotrexate treatment

Objectives:

We aimed to conduct the study "Research on cytokine changes and treatment effectiveness of generalized erythrodermic

psoriasis by Methotrexate " with the following objectives:

1 Investigate relevant factors and clinical characteristics of GEP patients undergoing treatment at the Central Dermatology Hospital

2 Determine the concentrations of cytokines: IL-2, IL-4, IL-6, IL-8, IL-10, IL-17, TNF-α, INF-γ in GEP patients before and after Methotrexate treatment

3 Evaluate the effectiveness of Methotrexate treatment for GEP

CHAPTER 1 DOCUMENTARY OVERVIEW 1.1 Generalized erythrodermic psoriasis

1.1.1 The situation of red psoriasis of the whole body

In Vietnam, in the National Hospital of Dermatology from 2008-2010, the rate of psoriasis patients coming to the clinic accounted for 2.2%, the proportion of psoriasis patients compared to inpatients accounted for about 20.1% According to Dang Van Em, patients with psoriasis hospitalized from 1990 to 1994 at the Department of Dermatology, Hospital 103 and Central Military Hospital 108 accounted for about 6.16% GEP was confirmed in psoriasis patients with skin lesions over 90% of the body surface In Vietnam, there are currently very few studies on systemic erythrodermic psoriasis

1.1.2 Pathophysiology of erythrodermic psoriasis

The pathogenesis of GEP is not fully understood, systemic erythrodermic psoriasis is a rare and severe variant of common psoriasis, however, some studies suggested this disease was associated primarily with the T helper 2 (Th2) phenotype Th2 cytokines (IL-13, IL-5, IL-10, IL-9) may be involved in the pathogenesis of GEP This change in cytokines causes normal psoriasis to turn into GEP Th1/Th2 imbalance is also the cause of GEP Understanding this pathogenesis helps scientists to find ways

to treat the disease such as using biological drugs against T cells, tumor necrosis factor-α (TNF-α), IL-12 axis /IL-23, or redirect cellular immune responses to the protective Th2 form of IL-4…

1.1.3 Clinical features of erythrodermic psoriasis

GEP is psoriasis diagnosed when the lesion area is greater than 90% of the body surface On the red skin, there are thick, infiltrated, edematous skin scales with different degrees depending on the location of the body and the severity of the disease GEP is a severe form of psoriasis and most of the time, it is converted from normal

psoriasis due to the treatment process of abuse of many drugs, especially systemic corticosteroids Many patients present with fever, chills, skin pain, extreme fatigue, tachycardia, diarrhea, constipation, weight loss, left heart failure due to excessive dehydration and edema, and visual disturbances GEP include two clinical forms, dry and wet

1.1.4 Treatment of erythrodermic psoriasis

The treatment strategy for psoriasis in general and GEP in particular has 2 phases: attack (reduce or clear the psoriasis lesions) and maintenance phase (maintain the psoriasis stability of the advanced stage) public), with the drug strategy: alone, in combination, in rotation and successively, has helped psoriasis patients in general and GEP patients have a good time of disease stabilization, improving their quality of life

Topical treatment drugs

+ Corticosteroids: For GEP, the treatment of corticosteroids needs

to be strictly indicated, and often combined with moisturizers to limit dry skin and limit systemic side effects of long-term topical corticosteroids

+Vitamin D: Vitamin D3 has 3 types: calcitriol (1,25(OH)2D3,

calcipotriol (MC903), a synthetic analogue of vitamin D with strong regulatory properties, very little risk of calcium and tacalciol changes

(1,24(OH)2D3)

Whole body medicine

+ Methotrexate: MTX is the preferred systemic drug for systemic erythrodermic psoriasis (details section 3.3)

+ Retinoid and derivatives: a synthetic derivative of vitamin A

acid, which regulates proliferation and differentiation of

keratinocytes

+ Cyclosporine A (CyA): Immunosuppressive, acting on many cell

types, preventing activation of Th, reducing INF-γ will cut off the

exchange between T-lymphocytes and T-cells

Mycophenolate mofetil: another immunosuppressive drug that

selectively inhibits activated lymphocytes that may be effective as

monotherapy for moderate to severe psoriasis

+ Probiotics: Several types of biologics have been used to treat

GEP, including TNF-α inhibitors, IL-12/IL-23 inhibitors, and most

recently inhibitors IL-17A

1.2 Role of cytokines in systemic erythrodermic psoriasis

Cytokines are low-molecular-weight soluble protein molecules produced by many cells in response to allergens that act as messengers to regulate inflammatory and immune function IL-4 and IL-10 are representative of Th2 cytokines IL-4 can promote B-lymphocyte activation, proliferation, synthesis, and secretion of IgE antibodies, and induce inflammatory responses by circulating immune complex accumulation on blood vessel walls IL-10 is produced by activated macrophages and some keratinocyte activators, and it can inhibit the production of IL-2, IFN-γ, proinflammatory cytokines and APC antigen may promote development of Th2 cytokines

1.3 Methotrexate for the treatment of erythrodermic psoriasis 1.3.1 Role of methotrexate in systemic erythrodermic psoriasis

Although there are now many biological drug classes that affect the pathogenesis of psoriasis, bringing positive effects in the stages of attack, maintenance and improvement of the quality of life for patients However, to date, Methotrexate (MTX) has been identified as a highly effective drug in the treatment of psoriasis in general

1.3.2 Studies on the treatment of systemic erythrodermic

psoriasis with methotrexate

Studies in the world: The study of Collins and Rogers made 7/40

patients with systemic erythrodermic psoriasis, treated with MTX at

a dose of 10mg/week, with 4 very good results, 2 patients with 2 patients well, 1 patient did not respond Van Dooren-Greebe studied

10 patients, with a dose of MTX 7.7-15mg/week, with 9 good results,

1 average results Haustein and Rytte had 36 patients, treated with MTX dose of 7.5-40mg/week then maintained at 7.5-15mg, 28 patients had good results, 6 patients had average results

Research in Vietnam: In Vietnam, there are many studies on the use

of Methotrexate in common psoriasis, but so far there is no study on the use of Methotrexate in erythrodermic psoriasis whole body skin

CHAPTER 2 SUBJECT AND RESEARCH METHODS 2.1 Subject and Research Materials

2.1.1 Research Subjects

Objective 1: 112 patients diagnosed with generalized erythrodermic

psoriasis (GEP) treated as inpatients and outpatients at the Central Dermatology Hospital from 01/01/2017 to 30/06/2019

Objective 2: 30 GEP patients (study group - SG) without

contraindications to use MTX and 30 healthy individuals (control group - CG) matched for age and gender, without autoimmune and infectious diseases

Objective 3: 30 GEP patients from SG in objective 2

2.1.1.1 Diagnostic criteria: Basic lesions: Red, scaly plaques

covering ≥90% of the patient's body surface area: History of

psoriasis vulgaris

2.1.1.2 Inclusion criteria

Objective 1: All patients diagnosed with GEP treated as inpatients

and outpatients at the Central Dermatology Hospital Patients agree

to participate in the study

Objective 2:

- Study group: 30 patients - GEP patients from objective 1, aged over

12, without contraindications to Methotrexate, and agree to participate in the study

- Control group: 30 healthy individuals, matched for age and gender with the patient group, without autoimmune or acute infectious diseases

Objective 3: Same as the study group in objective 2

- Control group: Not complying with the study protocol

Objective 3: Same as the study group in objective 2

2.1.2 Research materials:

- Methotrexate tablets: 2.5mg per tablet Supplied by the Pharmacy

Department of the Central Dermatology Hospital, manufactured in South Korea

- Reagents: Two kits and reagents for testing 7 cytokines (IL-2, IL-4,

IL-6, IL-8, IL-10, INF-γ, TNF-α) from Bio-Rad (USA) and testing kits for IL-17 and IL-23 from Sigma (USA)

+ Reagent kit for testing 7 cytokines (IL-2, IL-4, IL-6, IL-8, IL-10, TNF-α, and INF-γ) from Bio-Rad (USA)

+ A mixture of equal quantities of different types of plastic particles, each type attached to a different microplate, especially for cytokines that stimulate colony formation of mononuclear and granulocyte cells, interferon gamma (INF-γ), and tumor necrosis factor alpha (TNF-α)

+ Detecting antibody mixtures containing monoclonal antibodies specific to biotin-bound cytokines

+ Streptavidin-Phycoerythrin (PE) conjugate

+ Standard mixture of 27 human cytokines with known hormones + Sample solutions, biological solutions, washing solutions, running solutions produced and provided by Bio-Rad

+ Bio-Plex system and accompanying control software manufactured

by Rad company

+ Other auxiliary laboratory materials and equipment such as shakers, vacuum machines, pipettes, pipette tips, foil, blotting paper, distilled water, test tubes all meet international standards and are supplied from genuine suppliers manufacture

+ Biological chemicals are managed at the Department of Immunology-Military Medical Student

2.2 Research Methods

2.2.1 Research Design

- Objective 1: Prospective, Cross-sectional study

- Objective 2: Prospective, Cross-sectional study with a comparative control group

- Objective 3: Prospective controlled interventional trials

2.2.2 Research Sample Size

- Objective 1: Convenient sample - All GEP patients treated as inpatients and outpatients at the Central Dermatology Hospital, receiving direct examinations

- Objectives 2 and 3: Convenient sample (select GEP patients meeting the inclusion criteria for the study) and the final sample size must be ≥30 patients

2.2.3 Assessment of Disease Severity

Based on the Psoriasis Area & Severity Index (PASI) scale: Mild: PASI < 10; Moderate: 10 ≤ PASI < 20; Severe: PASI ≥ 20

2.2.4 Treatment Outcome Evaluation

- Clinical results are calculated by the percentage reduction in PASI according to the formula by Heng-Leong Chan-1993

- PASI reduction levels: Excellent: PASI reduction 100%; Good: 75

≤ PASI reduction < 100%; Fair: 50 ≤ PASI reduction < 75%;

Moderate: 25 ≤ PASI reduction < 50%; Poor, no effect: PASI reduction < 25%

2.4 Data Processing Method

- Data input and analysis using Epi InfoTM7 software Data presented in frequencies, percentages, mean values, standard

deviations, and medians

- Use the chi-square test to find associations for categorical variables or Fisher's exact test when there are >20% expected frequencies in a table <5, calculate OR with 95% confidence interval, and analyze variance using ANOVA Compare mean values for quantitative variables with normal distribution using paired T-test, and ANOVA to compare multiple mean values For non-normal distributions, use the non-parametric Wilcoxon test to compare two mean values and Kruskal-Wallis to compare multiple mean values

2.5 Location and Time of Study:

- Location: Central Dermatology Hospital; Department of

Immunology, Military Medical Academy

- Study period: From 01/01/2017 to 30/06/2019

CHAPTER 3 RESULTS 3.1 Some related factors, clinical characteristics of generalized erythrodermic psoriasis (GEP)

3.1.1 Some related factors

This study included 112 patients, in which the number of men was 97 and the number of women was 15, the male/female ratio was 6.5 The difference was statistically significant with p<0.01 Age

of onset <40 accounted for 58.93%, age of onset ≥40 accounted for 56.5% The age group 50-59 accounted for the highest percentage at 31.2%, followed by the age group over 60 accounted for 29.5%, the age group 30-39 accounted for 12.5%

The disease duration from 2-5 years was 16.1%, from 5-10 years was 32.14%, the highest rate was the group >10 years with the rate of

41.05, the group ≤1 year old accounted for 10, 71% The difference between groups was not statistically significant p>0.05

The time for patients with psoriasis to change to generalized erythrodermic psoriasis from 2-5 years accounted for the highest 49.12% The duration of illness ≤1 year accounted for 25.9%, the duration of illness from 5-10 years accounted for 22.32%, the duration of illness >10 years accounted for 2.66%, there were only 3 cases

Table 3.5 Predisposing factors associated to GEP (n=112)

Psychological trauma (Stress) 75 66,97

Localized bacterial infection 62 55,36

The symptoms of itching were 100 times, accounting for 89.29%, burning accounted for 34.82%, fatigue accounted for 38.39%, joint pain accounted for 27.68%, fever accounted for 11.61%

Nail damage: color change symptoms accounted for 77.68%, nail

fungus accounted for 52.68%, and at least nail pitting accounted for

only 6.25%

Location of disease onset: the most common site of disease onset is

the head area, accounting for 85.71%, followed by the upper

extremities 13.39% and at least the lower extremities 0.89%

Distribution according to PASI

Table 3.13 Distribution of PASI levels of patients with GEP (n=30)

3.2 Results of concentrations of 2, 4, 6, 8, 10,

IL-17, TNF-α, INF-γ of RTIs patients treated with Methotrexate 3.2.1 Subject characteristics of the study group and the control group

Table 3.14 Subject characteristics of the 2 groups

-Men

-Women

25 (83,3%) 5(16,7%)

24(80,0%)

6 (20,00%)

0,136 (Z-Test) The age and sex of the participants and the participants were similar with p>0.05 The mean PASI was 43.3±5.14

3.2.2 Results of IL-2, IL-4, IL-6, IL-8, IL-10, IL-17, TNF-α,

INF-γ concentrations of GEP patients treated with Methotrexate before treatment

Table 3.15 Comparison of the levels of cytokines before treatment of

the 2 groups

Cytokine

Patient group (n=30)