Targeting therapy in pemphigus: Where are we now and where are we going?

Pemphigus is a heterogeneous group of autoimmune skin disorders characterized by blistering of the skin and mucosal membranes, potentially affecting the quality of life if left unchecked. The current mainstay of treatment is systemic corticosteroids and immunosuppressive agents. Nevertheless, long-term use of these drugs can easily cause infections and other life-threatening adverse reactions. Thus, currently, researchers are trying to develop new and safer therapeutic approaches. Specifically, targeted therapies to pathogenic immune pathways have been gradually introduced and used for the treatment of pemphigus or in clinical trials, such as monoclonal anti-CD20 antibody, BAFF inhibitor, BTK inhibitor, CAAR-T therapy, FcRn antagonist, and TNF-α inhibitor. In addition, IL-4Rα antibody, IL-17 blockade, mTOR pathway inhibitor, CTLA-4Ig, and p38 MAPK inhibitors are theoretically promising treatment for pemphigus. Here, we review the research progress on the mechanism of targeted therapies for pemphigus.

Interleukin-37 inhibits desmoglein-3 endocytosis and keratinocyte dissociation via upregulation of Caveolin-1 and inhibition of the STAT3 pathway.

BACKGROUND: Pemphigus vulgaris (PV) is a potentially fatal autoimmune bullous disease primarily caused by acantholysis of keratinocytes attributed to pathogenic desmoglein-3 (Dsg3) autoantibodies. Interleukin-37 (IL-37) reportedly plays important roles in a variety of autoimmune diseases, but its role in PV is not clear. OBJECTIVES: To investigate whether IL-37 plays a role in the occurrence and progression of PV. METHODS: HaCaT keratinocytes were stimulated with anti-Dsg3 antibody to establish an in vitro PV model, which was defined as anti-Dsg3 group. Cells incubated with medium without anti-Dsg3 treatment were used as control. IL-37 was cultured with these cells infected with or without lentiviral vector shRNA-Caveolin-1 (sh-Cav-1-LV). Cell dissociation assay and immunocytofluorescence were performed to assess keratinocyte dissociation, keratin retraction and Dsg3 endocytosis. Real-time PCR was used to detect the mRNA level of Cav-1, and western blot was used to determine the protein expression of Cav-1, Dsg3, STAT3 and phosphorylated-STAT3 (p-STAT3). RESULTS: The anti-Dsg3 group showed more cell debris, increased keratin retraction, increased Dsg3 endocytosis, reduced Cav-1 expression and co-localization than the control group, while IL-37 treatment neutralized all of these changes. Interestingly, Cav-1 knockdown supressed the inhibitory effect of IL-37 on keratinocyte dissociation and Dsg3 internalization. The protein expression of p-STAT3 was increased in keratinocytes of the PV model but decreased by IL-37. Re-activation of the STAT3 pathway by colivelin supressed the inhibitory effect of IL-37 on keratinocyte dissociation and Dsg3 internalization, along with upregulation of Cav-1 and Dsg3. CONCLUSIONS: IL-37 inhibited keratinocyte dissociation and Dsg3 endocytosis in an in vitro PV model through the upregulating Cav-1 and inhibiting STAT3 pathway.

Nine cases of refractory bullous pemphigoid treated with dupilumab and literature review.

BACKGROUND AND AIMS: Bullous pemphigoid is an autoimmune blistering disease that affects the elderly mostly. First-line treatment of systemic corticosteroids may cause significant adverse effects, especially in patients with multiple co-morbidities. Dupilumab shows certain effectiveness in treating BP. We aim to profile our experience with dupilumab in a series of patients with BP and review the articles published to date. METHODS: Medical records of 9 patients with moderate-to-severe BP were retrospectively reviewed. All patients were administered dupilumab. Response to dupilumab was evaluated by NRS scores, number of lesions, and the systemic corticosteroids' dosage. The PubMed, Embase, and Web of Science databases were searched to identify eligible studies. RESULTS: The 9 patients were identified in this case series with a median age of 68 years (range 42-89) and the median duration of disease before being treated with dupilumab was 6 months (range 1-144). Complete remission was achieved in 6 patients while partial response was achieved in one patient. The NRS score had decreased to varying degrees at week 2 in all patients, and skin lesions improved within 2 to 6 weeks. Fifteen publications were included: 3 retrospective studies and 12 case series or reports, with a total of 63 patients. The overall complete response and partial response rates were 74.6 % and 11.1 %, respectively. CONCLUSION: Dupilumab appears to be a safe alternative for the treatment of patients with refractory BP.

Systematic review and network meta-analysis of different types of emollient for the prevention of atopic dermatitis in infants.

Prophylactic application of emollients has been an effective strategy against infant atopic dermatitis (AD); however, the difference of different emollients is unknown. We performed this network meta-analysis to compare different emollients in preventing infant AD. A systematic search was performed in PubMed, EMBASE and Cochrane library to identify relevant studies from their inception through 28 February, 2022. We evaluated the quality of eligible studies using the Cochrane risk of bias assessment tool. Data analysis was performed using STATA 14.0. Eleven studies were included for data analysis. Direct meta-analysis suggested that early application of emollients effectively prevented AD development in high-risk infants (risk ratio [RR], 0.64; 95% confidence interval [CI], 0.47 to 0.88). Network meta-analysis suggested that emollient emulsion might the better option for preventing infant AD development, with a surface under the cumulative ranking curve (SUCRA) of 82.6% for all populations, 78.0% for high-risk populations and 79.2% for populations with food sensitization. Moreover, subjects receiving emollients more frequently experienced adverse events. Overall, early application of emollients is an effective strategy for preventing AD development in high-risk infants and emollient emulsion may be the optimal type. Future study with well-designed and large scale are warranted to validate our findings.

UVB-induced SFRP1 methylation potentiates skin damage by promoting cell apoptosis and DNA damage.

In response to the accumulation of genetic mutations and cellular changes, ultraviolet radiation B (UVB) skin lesions undergo dysplasia and transform into a cutaneous squamous cell carcinoma (CSCC). Consistent with our previous findings that secreted frizzled-related protein 1 (SFRP1), a member of the SFRP gene family, was downregulated in human CSCC tissue samples, we found a significant downregulation of SFRP1 in HaCaT, A431, and SCL-1 cells after UVB irradiation. DNA methyltransferase 1 (DNMT1) was significantly increased in CSCC tissues as well as UVB-exposed A431 and SCL-1 cells. Bisulfite genomic sequencing analysis showed that the downregulation of SFRP1 was mainly due to methylation of the SFRP1 promoter, as indicated by increased methylation rate of SFRP1 after UVB irradiation in HaCaT cells. Moreover, demethylation treatment with 5-aza'-deoxycytidine (5-AzaC) increased SFRP1 expression and reduced the methylation rate of SFRP1 in HaCaT cells. Flow cytometry analyses demonstrated that 5-AzaC treatment or overexpression of SFRP1 ameliorated UVB-induced apoptosis, while knockdown of SFRP1 promoted UVB-induced apoptosis in HaCaT cells. In addition, a comet assay confirmed that 5-AzaC treatment reduced DNA damage following UVB irradiation, while knockdown of SFRP1 enhanced DNA damage following UVB irradiation. In conclusion, our study identified DNA methylation of SFRP1 as a key mediator in the UVB-induced apoptosis of keratinocytes. These findings indicate that reinforcing SFRP1 defences by 5-AzaC may help prevent UVB-induced skin damage.

Safety and Efficacy of Nemolizumab for Atopic Dermatitis With Pruritus: A Systematic Review and Meta-Regression Analysis of Randomized Controlled Trials.

Background: Nemolizumab is deemed as a promising drug for atopic dermatitis (AD) patients with pruritus. Objective: This study aimed to evaluate the efficacy of nemolizumab in treating patients with AD and the association between the dosage or regimen of nemolizumab with the improvement in clinical indices. Methods and Materials: PubMed, Embase, and the Cochrane Library were searched for randomized controlled trials (RCTs) published from inception to August 2021. Results: A total of 14 cohorts of participants from six randomized controlled studies were included in the meta-analysis. Nemolizumab significantly reduced the pruritus VAS (WMD = -18.86, 95% CI: -27.57 to -10.15, p < 0.001; I2 = 56.2%, pheterogeneity = 0.005) and EASI (WMD = -11.76, 95% CI: -20.55 to -2.96, p = 0.009; I2 = 0%, pheterogeneity = 0.978) scores compared with placebo. No significant difference was observed in the occurrence of any AEs (RR = 1.03, 95% CI: 0.93 to 1.13, p = 0.593; I2 = 0%, pheterogeneity = 0.980) between the two groups. The univariate meta-regression showed that both the dosage and study duration had no association with the change of pruritus VAS score. Conclusion: Nemolizumab presented a promising effect based on the difference in the average change in pruritus VAS and EASI scores compared with placebo. The results indicated its efficacy in relieving pruritus and the severity of AD and improving patients' quality of life.

Mechanism underlying the effect of SO2-induced oxidation on human skin keratinocytes.

This study aimed to study the effect and mechanism of action of SO2-induced oxidation on human skin keratinocytes.Different concentrations of SO2 derivatives (0, 25, 50, 100, 200, 400, and 800 µM) were used for treating HaCaT keratinocytes for 24 hours. MTT was used to evaluate the effect of each concentration on cell proliferation. HaCaT cells were randomly divided into control and SO2 groups. The control group received no treatment, whereas the SO2 group was treated with SO2 derivatives of selected concentrations for 24 hours. The levels of reactive oxygen species (ROS), malondialdehyde (MDA), and superoxide dismutase (SOD), tumor necrosis factor TNF-α (TNF-α), and interleukin-1 (IL-1-ß) in cell supernatants were detected using enzyme-linked immunosorbent assay. Real-time polymerase chain reaction was used to detect the expression of nuclear transcription factor (Nrf2) and heme oxygenase (HO)-1 mRNA. The Western blot analysis was used to test the expression levels of Nrf2, HO-1, activated caspase-3, Bcl-2, Bax, IκB, NF-κB p65 (p65), ERK1/2, p38, phospho-NF-κB p65 (p-p65), p-ERK1/2, and p-p38.SO2 derivatives (100, 200, 400, and 800 µM) could inhibit cell proliferation. SO2 derivatives increased the level of ROS, MDA, TNF-α, IL-1ß, Nrf2, HO-1, and p-p65/p65 and decreased the levels of SOD, IκB, p-ERK1/2/ERK1/2, and p-p38/p38 compared with the control group, but they had no effect on the levels of caspase-3, Bcl-2, and Bax.SO2 could inhibit the proliferation of human skin keratinocytes and induce oxidative stress and inflammation via the activation of the NF-κB pathway to inhibit the ERK1/2 and p38 pathways.

Identification of key genes and pathways involved in vitiligo development based on integrated analysis.

Vitiligo is a chronic skin condition lack of melanocytes. However, researches on the aetiology and pathogenesis of vitiligo are still under debate. This study aimed to explore the key genes and pathways associated with occurrence and development of vitiligo.Weighted gene coexpression network analysis (WGCNA) was applied to reanalyze the gene expression dataset GSE65127 systematically. Functional enrichments of these modules were carried out at gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), gene set variation analysis (GSVA), and gene set enrichment analysis (GSEA). Then, a map of regulatory network was delineated according to pivot analysis and drug prediction. In addition, hub genes and crucial pathways were validated by an independent dataset GSE75819. The expressions of hub genes in modules were also tested by quantitative real-time polymerase chain reaction (qRT-PCR).Eight coexpressed modules were identified by WGCNA based on 5794 differentially expressed genes of vitiligo. Three modules were found to be significantly correlated with Lesional, Peri-Lesional, and Non-Lesional, respectively. The persistent maladjusted genes included 269 upregulated genes and 82 downregulated genes. The enrichments showed module genes were implicated in immune response, p53 signaling pathway, etc. According to GSEA and GSVA, dysregulated pathways were activated incessantly from Non-Lesional to Peri-Lesional and then to Lesional, 4 of which were verified by an independent dataset GSE75819. Finally, 42 transcription factors and 228 drugs were spotted. Focusing on the persistent maladjusted genes, a map of regulatory network was delineated. Hub genes (CACTIN, DCTN1, GPR143, HADH, MRPL47, NKTR, NUF2) and transcription factors (ITGAV, SYK, PDPK1) were validated by an independent dataset GSE75819. In addition, hub genes (CACTIN, DCTN1, GPR143, MRPL47, NKTR) were also confirmed by qRT-PCR.The present study, at least, might provide an integrated and in-depth insight for exploring the underlying mechanism of vitiligo and predicting potential diagnostic biomarkers and therapeutic targets.

The association between rs2476601 polymorphism in PTPN22 gene and risk of alopecia areata: A meta-analysis of case-control studies.

BACKGROUND: The single nucleotide polymorphism (SNP) rs2476601 of the protein tyrosine phosphatase, nonreceptor type 22 (PTPN22) gene has been presented to implicate in the pathogenesis of alopecia areata (AA) in a few association investigations with limited sample size and inconsistent conclusions. METHODS: The aim of the current meta-analysis was to assess and synthesize the presently available data on the connection between rs2476601 and AA vulnerability. Six electronic databases, including EMBASE, PubMed, Web of Science, the Cochrane Library, Wanfang data, and the China National Knowledge Infrastructure database (CNKI), were systematically retrieved for relevant observational studies published previous to November 2018. Total odds ratios (ORs) and corresponding 95% confidence intervals (95% CIs) were analyzed to evaluate the correlation between PTPN22 polymorphism and AA. Risk of bias was estimated according to the Newcastle-Ottawa Scale (NOS). Sensitivity analyses were carried out using the RevMan 5.3 software. RESULTS: In general, 5 case-control studies including 1129 AA patients and 1702 healthy control individuals were obtained for this meta-analysis. The pooled results suggested that rs2476601 SNP was significantly associated with AA susceptibility under allelic model (C vs T, OR = 0.77, 95% CI, 0.64-0.92, P = .003) and recessive model (CC vs CT + TT, OR = 0.73, 95% CI, 0.60-0.88, P = .001). CONCLUSION: On the basis of the results of the current research, the rs2476601 polymorphism of PTPN22 gene is significantly correlated with AA susceptibility. The C-allele and CC-genotype carriers at this locus have a lower risk of AA.

MiR-497-5p, miR-195-5p and miR-455-3p function as tumor suppressors by targeting hTERT in melanoma A375 cells.

BACKGROUND: hTERT gene plays an important role in melanoma, although the specific mechanism involved is unclear. The aim of this study was to screen and identify the relative miRNAs with the regulation of hTERT in melanoma. MATERIALS AND METHODS: Quantitative real-time polymerase chain reaction (q-PCR) and immunohistochemistry were performed to detect hTERT mRNA and protein expression in 36 formalin-fixed paraffin-embedded melanoma tissues and 36 age- and sex-matched pigmented nevi cases, respectively. Bioinformatics analysis and custom miRNA polymerase chain reaction array were determined for predicting, screening and verifying miRNAs with the regulation of the hTERT gene. To investigate the biological functions, miRNAs mimics or inhibitors were transfected into melanoma A375 cells. The relative expression of miR-497-5p, miR-195-5p, miR-455-3p and hTERT mRNA was determined by q-PCR. The protein expression of hTERT was detected by Western blot. 3-(4,5-Dimethylthiazolyl-2-yl)-2,5-biphenyl tetrazolium bromide and flow cytometry were employed to detect cell proliferation ability, cell apoptosis and cell cycle. Transwell and wound healing assays were used to observe cell invasion and migration abilities. A direct target gene of miRNAs was analyzed by a dual luciferase reporter activity assay. RESULTS: MiR-497-5p, miR-195-5p, miR-455-3p were significantly downregulated, while hTERT was upregulated in melanoma tissues. hTERT expression level was inversely correlated with miR-497-5p, miR-195-5p and miR-455-3p. Overexpression of miR-497-5p, miR-195-5p and miR-455-3p inhibited A375 cell proliferation, migration and invasion, arrested the cell cycle, induced cell apoptosis and decreased hTERT expression at both mRNA and protein levels. Suppression of miR-497-5p, miR-195-5p and miR-455-3p partially reversed the inhibitory effects. Finally, hTERT was identified as a direct target of miR-497-5p, miR-195-5p and miR-455-3p. CONCLUSIONS: MiR-497-5p, miR-195-5p and miR-455-3p act as tumor suppressors by targeting hTERT in melanoma A375 cells. Therefore, miR-497-5p, miR-195-5p and miR-455-3p could be potential targeted therapeutic choice for melanoma.

Aberrations and clinical significance of BRAF in malignant melanoma: A series of 60 cases in Chinese Uyghur.

Malignant melanoma (MM) is a highly malignant melanocytic tumor, it occurs mostly in the skin, the mucous membrane close to the skin, but also in the tunicae rhagoides and the pia mater. The Uyghur is the largest ethnic group living in the Xinjiang Uyghur Autonomous Region of China, accounting for 46% of the total population of 20 million. Large-scale studies on MMs in Asian countries are limited. This study aimed to investigate BRAF mRNA expression and mutations in Chinese Uyghur patients with MMs and to identify the clinical features associated with these parameters.Formalin-fixed, paraffin wax-embedded tumor sections from 60 MMs were analyzed for BRAF expression using reverse transcription polymerase chain reaction (RT-PCR). Exons 11 and 15 of BRAF were analyzed for the presence of mutations using PCR and DNA sequencing. Sixty MMs were followed by mobile phone for survival analysis.BRAF mRNA expression was higher in MMs than in pigmented moles and normal skin tissues. Fourteen of 60 MMs had BRAF mutations. The frequency of BRAF mutations was significantly higher in patients younger than 60 years (10/28, 4/32, P = .02). A significant difference was observed in the frequency of BRAF mutations among specimens of mucosal, acral, chronic sun-induced damage (CSD), and non-CSD MMs (2/10, 3/19, 8/25, 1/6, P = .002). No significant association was found among BRAF mutations, sex, ulceration, or lymph node metastasis. MMs lymph node metastasis (hazard ratio 2.54 [95% confidence interval 1.062 - 6.066], P = .01) affected survival.This study indicated that BRAF mutations and expression might serve as independent adverse prognostic factors in melanoma.

Naringenin protects keratinocytes from oxidative stress injury via inhibition of the NOD2-mediated NF-κB pathway in pemphigus vulgaris.

Naringenin is known to have anti-oxidative activity; however, the effect of naringenin on the progression of pemphigus vulgaris (PV) still remains unclear. This study aims to analyze the effect of naringenin on HaCaT cell apoptosis and oxidative damage under the treatment of PV serum. The results showed that PV serum significantly induced cell apoptosis compared with the control group; whereas, comparing with PV group, naringenin inhibited cell apoptosis. Moreover, PV serum increased the expression of bax and caspase-3, and decreased the expression of bcl-2; but naringenin significantly suppressed the expression of bax and caspase-3, induced the expression of bcl-2. Naringenin inhibited PV serum-induced disruption of cell-cell contacts. Naringenin also down-regulated the expression of Dsg1, Dsg3 and E-cadherin compared with the PV group. Additionally, naringenin noticeably decreased the PV serum-induced ROS production and alleviated PV serum induced the drop of mitochondrial membrane potential. Furthermore, naringenin increased the activity of SOD, GSH-Px and TAC under the treatment of PV serum. Naringenin also decreased the expression of NOD2, RIPK2 and NF-κB p-p65, but this effect could be reversed by muramyl dipeptide (MDP). In conclusion, these results suggested that naringenin protected keratinocytes from apoptosis and oxidative stress injury through inhibition of the NOD2-mediated NF-κB pathway.

Blocking RhoA/ROCK inhibits the pathogenesis of pemphigus vulgaris by suppressing oxidative stress and apoptosis through TAK1/NOD2-mediated NF-κB pathway.

Oxidative stress and apoptosis play critical roles in pemphigus vulgaris (PV). The main aim of the present study was to investigate the effects of RhoA/ROCK signaling on UVB-induced oxidative damage, and to delineate the molecular mechanisms involved in the UVB-mediated inflammatory and apoptotic response. In HaCaT cells, we observed that blockage of RhoA/ROCK signaling with the inhibitor CT04 or Y27632 greatly inhibited the UVB-mediated increase in intracellular reactive oxygen species (ROS). Additionally, inhibition of RhoA/ROCK signaling reduced UVB-induced apoptosis, as exemplified by a reduction in DNA fragmentation, and also elevated anti-apoptotic Bcl-2 protein, concomitant with reduced levels of pro-apoptotic protein Bax, caspase-3 cleavage and decreased PARP-1 protein. The release of inflammatory mediators TNF-α, IL-1ß, and IL-6 was also attenuated. Mechanically, we observed that blockage of RhoA/ROCK repressed the TAK1/NOD2-mediated NF-κB pathway in HaCaT cells exposed to UVB. Taken together, these data reveal that RhoA/ROCK signaling is one of the regulators contributing to oxidative damage and apoptosis in human keratinocytes, suggesting that RhoA/ROCK signaling has strong potential to be used as a useful therapeutic target in skin diseases including PV.

Secreted frizzled-related protein promotors are hypermethylated in cutaneous squamous carcinoma compared with normal epidermis.

BACKGROUND: The Wnt signaling pathway is abnormally activated in many human cancers. Secreted frizzled-related proteins (SFRPs) function as negative regulators of Wnt signaling and play an important role in carcinogenesis. SFRP promoter hypermethylation has often been identified in human cancers; however, the precise role of SFRPs in cutaneous squamous cell carcinoma (SCC) is unclear. METHODS: The methylation status of the SFRP family was analyzed in an age-and sex-matched case-control study, including 40 cutaneous SCC cases and 40 normal controls, using the MassARRAY EpiTYPER system. RESULTS: The methylation rate of SFRP1, SFRP2, SFRP4, and SFRP5 promoters was significantly higher in cutaneous SCC tissues than in adjacent tissue and normal skin samples. DISCUSSION: Our manuscript mainly discussed the average methylation rate of SFRPs (SFRP1, SFRP2, SFRP4, and SFRP5) promoters are significantly high in tumor tissue samples and the average CpG island methylation rate among different pathological levels of cutaneous SCC between these genes are different. CONCLUSIONS: Our findings suggest that promoter hypermethylation of SFRPs is associated with the development of carcinoma, and could be a useful tumor marker for cutaneous SCC and other types of cancers.