Genetics and Epigenetics of Keloids

Abstract

Significance:

Keloid scarring is cosmetically disfiguring, psychosocially distressing, and can be physically disabling. The pathophysiology of keloid formation is poorly understood and subsequently, treatment options are ill defined, limited, and largely unsatisfactory. Therefore, in view of its unsatisfactory and recalcitrant management, keloid therapy is often seen as a financial burden affecting both patients and the health care systems.

Recent Advances:

Increased research on the genetic and epigenetic mechanisms in keloids has broadened our understanding of keloid pathobiology. Epigenetic mechanisms, mainly DNA methylation, histone modification, and noncoding RNAs, are currently being widely investigated. Advances in genetic sequencing technology and reduced cost have aided this endeavor. Studies on blood and patient-derived keloid tissue are being done with therapeutic agents targeting epigenetic and genetic markers with the shared goal of identifying the pathways underlying the initiation and maintenance of keloids. These advances have informed us of multiple complex molecular pathways implicated in keloids, which are yet to be fully elucidated.

Critical Issues:

Improved understanding of the genetic and epigenetic causes implicated in keloids will enhance our knowledge of this enigmatic disorder and likely lead to the development of therapeutic targets based on the available clinical and experimental studies. Due to the incomplete knowledge of molecular targets involved in keloid scarring pathways, therapeutics is still lagging for this clinically and scientifically important condition.

Future Directions:

Focused research on the identification of molecular targets and mechanistic pathways implicated in keloids is required to generate novel antifibrotic therapeutic options to decrease or eradicate recurrence of the disease as well as associated morbidity and improve the quality of life of those affected with keloids.

Ardeshir Bayat, BSc (Hons), MBBS, PhD

Scope and Significance

A keloid is a benign hyperproliferative growth of dense fibrous tissue developing from an abnormal healing response to a cutaneous injury. Genetic, epigenetic, and environmental mechanisms contribute to keloid formation. Although some key cellular and molecular processes underlying keloid scarring are known, a complete understanding of keloid pathogenesis remains unclear on how these pathogenetic pathways are initiated and maintained. This is evidenced by the paucity of effective therapies and even fewer that target fibrogenesis specifically.

In this review, we summarize the genetic and epigenetic findings to date with examples that reveal specific solutions and potential challenges in keloid scarring.

Translational Relevance

Research in epigenetics has provided a better understanding of keloids, which was previously not well explained by genetics alone. Genetic and epigenetic mechanisms, whether singly or in tandem, have been shown to play a pivotal role in the induction and persistent activation of keloid fibroblasts. Not only have studies allowed us to better understand processes affecting the severity of disease forms but they have also allowed us to determine how they might contribute to a spectrum of clinical outcomes. This translational knowledge is key to the development of effective personalized therapeutic solutions in this area of clinically significant unmet need.

Clinical Relevance

Keloid scarring is often known to be pruritic, painful, disfiguring, and has a significant psychological impact on a patient's quality of life. It is a health concern as it is costly and has ill-defined unsatisfactory (often recurrent) treatment options. Multiple treatment modalities have been tried, but to date, there is no single effective therapy; hence, the condition remains a therapeutic challenge.

Overview

Keloids are proliferative fibrous growths resulting from an excessive response to cutaneous injury.¹ In distinction from a hypertrophic scar, keloids behave like benign tumors as they progressively continue to grow beyond the boundaries of the original wound margins without evidence of spontaneous regression and recur as they are resistant to treatment.²

The main effector cell is the fibroblast that produces excessive extracellular matrix components, particularly type 1 and 3 collagen.³ There are a wide variety of postulated triggers and disease-specific pathogenetic processes leading up to keloid scarring, involving the transforming growth factor β (TGF-β) family pathways, homologs of the drosophila protein, mothers against decapentaplegic and the C. elegans protein SMA (SMAD) (second messenger) proteins, p53, epidermal growth factor receptor (EGFR), and mammalian target of rapamycin (mTOR) signaling pathways, along with the wingless related integration site 5a (WNT5a)/β catenin signaling pathway.^3
–5

These are so expansive to preclude a detailed discussion, please see Fig. 1 for an overview of several important implicated molecular pathways and the effect of genetic and epigenetic modifications on some of these pathways. Nevertheless, the exact cause of the aberrant cellular and molecular mechanisms leading to excessive extracellular matrix accumulation in keloids largely remains unclear. Disease models are required to better understand keloid pathogenesis, yet it is not possible to establish keloids in animals because of the uniqueness of this disease to humans.

Figure 1.

Major signaling pathways and effects of epigenetic modifications on keloid scarring. Acute and chronic inflammation can trigger skin fibrosis in keloids. Examples of major pathways driving fibrosis are shown. After initiation of these pathways, (some through epigenetic modifications) fibroblasts are stimulated to proliferate and this is associated with reduced apoptosis, increased collagen production, increased cytokine production, and secretion of excess extracellular matrix proteins with concomitant reduced MMP production. EGFR, epidermal growth factor receptor; HOXA, homeobox A cluster; LINCO, long intergenic nonprotein coding RNA; lncRNA, long noncoding RNA; miRNA, microRNA; MMP, matrix metalloproteinases; mTOR, mammalian target of rapamycin; SFRP1, secreted frizzled-related protein 1; SMAD, homologs of the drosophila protein, mothers against decapentaplegic and the C. elegans protein SMA; TGFβ, transforming growth factor β; WNT, wingless related integration site. Color images are available online.

Some authors believe that keloids occur primarily in areas of tension.⁶ Having said that, keloids have a predilection for the presternum, deltoid, and suprapubic regions; however, the earlobe is also one of the most common anatomical sites affected by keloids and yet it is under minimal tension.⁶ Keloids can occur anywhere on the body, but rarely palms and soles where significant skin tension is also expected.²

Keloid scar formation can ultimately result in clinical and cosmetic sequelae with major concerns. Significant disfigurement, pruritus, and pain can also occur. Keloid scarring is a challenging health burden and concern to the clinician and patient as it remains recalcitrant to conventional therapy. Keloids present with high recurrence rates following most current treatment modalities.

Genetic and, more recently, epigenetic mechanisms and their interplay have been shown to partly account for the complexity of keloid scarring. This review article will specifically focus on our current understanding of keloids from a genetic and an epigenetic perspective, as well as discuss the potential of functional and translational application of therapeutic targets based on the available genetic and epigenetic studies.

Discussion

Genetics of keloid scarring

Familial keloids

The risk of developing keloids is 15 times more in dark-skinned individuals compared to whites.⁷ Keloid epidemiology data are limited, but the reported incidences in dark-skinned individuals and Hispanics range from 5% to 16%.² Most keloids occur sporadically, but some cases are familial.¹ Family survey data have suggested an autosomal dominant inheritance pattern with incomplete penetrance, although keloid transmission does not follow a simple Mendelian monogenic disease pattern, but appears to be rather polygenic or oligogenic.¹

Keloid scarring in monogenic disorders

Rare genetic disorders with a propensity for keloid scarring, including Dubowitz syndrome, Bethlem myopathy, Rubinstein-Taybi syndrome, Noonan syndrome, and Goeminne syndrome,⁸ have been reported. However, there is no one specific gene associated with the development of keloids, although several genes and gene loci have been implicated.

Genetic linkage analyses

A few keloid susceptibility loci and candidate genes have been identified using genome-wide linkage analysis (Table 1). This table includes a study with a logarithm of odds (LOD) less than 3 with a proposed gene implicated in keloid scarring⁹ and such information is valuable when dealing with different ethnic groups to point out locus heterogeneity in familial keloids.¹⁰

Table 1.

Loci identified through linkage analyses in keloids

Ethnicity	Locus	LOD Score	Proposed Gene	Reference
African American	7p11	3.16	EGFR	¹¹
Japanese	2q23	3.01	TNFAIP	¹¹
Chinese Han	18q21	2.201	SMAD2,4,7, PIAS2	⁹
Nigerian Yoruba	8p23.3-p21.3	4.48	ASAH1	¹²

ASAH1, N-acylsphingosine amidohydrolase; EGFR, epidermal growth factor receptor; LOD, logarithm of the odds; PIAS2, protein inhibitor of activated signal transducer and activator of transcription 2; SMAD, homologs of the drosophila protein, mothers against decapentaplegic and the C. elegans protein SMA; TNFAIP, tumor necrosis factor-alpha-induced protein.

Two unrelated large families were identified through probands in the United States and Japan. The first was an African American family, which had 13 affected members, and the other was a Japanese family with 7 affected members.

A genetic linkage study identified keloid susceptibility loci in the Japanese family on chromosome band 2q23 with a LOD score of 3.01 and the African American family on 7p11 with a LOD of 3.16, respectively, through peripheral blood samples.¹¹ The tumor necrosis factor-alpha-induced protein (TNFAIP) 6 gene at chromosome band 2q23 and the EGFR gene at chromosome band 7p11 were proposed to be the candidate genes within these respective loci.¹¹ TNFAIP6 is a gene involved in extracellular matrix stability and cell migration, and high TNFAIP6 protein expression is significantly related to aggressive pathological features in cancers.¹¹ A separate study with 32 affected members in a Chinese keloid pedigree with 5 affected generations, however, could not confirm linkage on chromosome 7p11, which, according to the authors, suggested locus heterogeneity in familial keloids.¹⁰ In the same Chinese Han pedigree, the linkage between two susceptibility loci,15q22.31-q23 and 18q21.1, to keloids, was investigated. This study suggested that keloid susceptibility may also be on chromosome 18q21 with LOD of 2.201.⁹

The 18q21 locus was proposed to contain SMAD 2, 7, and 4 genes as well as the protein inhibitor of activated signal transducer and activator of transcription 2 (PIAS2) gene. Of even more significance is that the SMAD genes are known to participate in the regulation of the TGF-β signaling pathway. The TGF-β pathway is important in virtually all types of fibrosis and is a potent stimulator of the synthesis of extracellular matrix proteins in most fibrogenic cells.³ The SMAD signal transduction pathway is a downstream mediator of TGF-β. Moreover, in keloids, TGF-β has been shown to act as a primary modulator of fibrosis, as exogenous TGF-β stimulates keloid fibroblast proliferation and collagen synthesis, while also inhibiting the collagen-degrading activity of matrix metalloproteinases (MMP).⁵

In an African study, a large Nigerian Yoruba family consisting of 24 family members with 9 affected individuals had a whole-genome analysis done on blood samples, which mapped a locus to chromosome 8p23.3-p21.3 with a maximum multipoint LOD score of 4.48. The proposed susceptibility gene was the N-acylsphingosine amidohydrolase (ASAH1) gene, although a clear mechanism could not be delineated.¹²

ASAH1 is an acid ceramidase known to be involved in apoptosis, cell cycle, differentiation, and cell invasion/tumor formation by controlling the ratio of ceramide and sphingosine, and is also expressed in cancer cell lines.¹² Since both TNFAIP6 and ASAH1 are implicated in cancers, it may be suggested that the quasi-neoplastic tendencies of keloids might be due to this effect of invading into adjacent normal tissue perpetuating keloid scarring.

Sporadic keloids

Human leukocyte antigen immunogenetics

As illustrated (Table 2), peripheral blood mononuclear cells showed an increased frequency of human leukocyte antigen (HLA) DR, DQ, and DP in keloid susceptible patients.¹³ In another study on keloid tissue, there was a positive association between HLA A*03, A*25, Cw*802, B*07, and A*01 in Chinese Han individuals,¹⁴ while in a separate study, also on keloid tissue, it gave differing results in that there was no association between keloid scarring and HLA A*01, A*03, A*25, B*07, Cw*802, DDQA1, and DQB1 in a Jamaican Afro-Caribbean ethnic group.¹⁵

Table 2.

Human leukocyte antigen immunogenetics of keloid susceptible patients

HLA	Increased/Decreased	Tissue/Cell/Blood	Population	References(s)
DR	↑	Peripheral blood mononuclear cells	Chinese	¹³
DQ	↑
DP	↑
A ^*03	↑	Tissue	Chinese	¹⁴
A ^*25	↑
Cw^*802	↑
B ^*07	↑
A ^*01	↑
A ^*01	No differences	Tissue	Jamaican Afro-Caribbean	¹⁵
A ^*03
A ^*25
B ^*07
Cw^*802
DDQA1
DQB1
DRB1^*15	↑	Tissue	Chinese/Caucasian	^16,17
DQA1^* 0104	↑	Tissue	Chinese	¹⁹
DQB1^* 0501	↑
DQB1^* 0503	↑
DQA1^* 0501	↓
DQB1^* 0201	↓
DQB1^* 0402	↓

HLA, human leukocyte antigen.

Two separate studies suggested an association of keloid susceptibility with HLA DRBI*15 in both Chinese Hans and Caucasians.^16,17 DRB1 increases susceptibility, not only in the Chinese but also in an Afro-Caribbean population.¹⁸

Another study, which analyzed HLA-DQA1 and HLA-DQB1 in Chinese Hans, found a positive association of HLADQA1*0104, DQB1*0501, and DQB1*0503, but a negative association of HLA-DQA1*0501, DQB1*0201, and DQB1*0402 with keloids compared with control subjects.¹⁹

Despite few similarities, the variation of the HLA immunogenetics between and within ethnicities shows that more research is still needed as we do not yet fully understand the role of immunogenetics in keloid scarring.

Genome-wide association studies

Multiple studies (Table 3), have identified keloid-related single-nucleotide polymorphisms (SNPs) through genome-wide association studies.^20
–22 Four SNPs (rs873549, rs1511412, rs940187, and rs8032158) in three chromosomal regions (1q41, 3q22.3–23, and 15q21.3) were found to be significantly associated with keloid disease in a Japanese population.²⁰ The locus 15q21.3 was found to have the neural precursor cell expressed developmentally downregulated protein (NEDD4) gene.

Table 3.

SNPs associated with keloids from genome-wide association studies

SNP	Locus	Ethnicity	Gene	Reference(s)
rs873549	1q41	Japanese/Chinese	—	^20,24
rs1511412	3q22.3–23	Japanese	—	²⁰
rs940187	3q23	Japanese	—	²⁰
Rs2271289	15q21.3	Chinese	NEDD4	²¹
Rs8032158	15q21.3	Japanese	NEDD4	^20,22
Rs747722	15q21.2–22.3	African American	MYO1E	^{25^a}
Rs28394564			MYO1E
Rs138585173			NEDD4
rs35641839	11q13.5	African American	MYO7A	^{25^a}

Admixture mapping.

MYO, myosin; NEDD, neural precursor cell expressed developmentally downregulated protein; SNP, single-nucleotide polymorphism.

A different study implicated a specific transcript variant of NEDD4, NEDD4-TV3, as a regulator of keloid formation by activating NF-kB/STAT3-mediated inflammation or by stimulating insulin-like growth factor (IGF)-1 signaling, or through other pathways as higher transcript levels of NEDD4- TV3 were noted in keloid keratinocytes and fibroblasts when compared with normal skin.²³ The locus1q41 and SNP rs873549 were later validated in a Chinese cohort.²⁴

An admixture mapping performed on an African American cohort identified SNPs within the myosin (MYO)1E gene, which is near NEDD4 on chromosome bands 15q21.2–22.3, as being associated with keloid occurrence.²⁵ This suggested there were common genetic elements on this locus. The same study also identified the SNP rs35641839 in the chromosomal band 11q13.5 within the MYO7A gene.

Several candidate genes, which may be involved in keloid scarring, have been investigated through mutations and polymorphism screening studies. The complex TGF-β pathway plays an important role in virtually all fibrosing diseases. This knowledge has prompted considerable research, which has focused on TGF-beta signaling crosstalk, especially TGF-β1, 2, and 3. Interestingly, a study in a Caucasian population demonstrated a lack of association between the TGF-β3 gene and keloidal scarring.²⁶ A separate study done on select SMAD gene SNPs in a Jamaican population showed no strong association with an increased risk of developing keloids, although this could not be completely ruled out.²⁷

Microarray studies

Even though several microarray studies have been conducted to ascertain the association between certain genes and keloid scarring, the reported dysregulated genes tend to vary. A lack of accuracy and reproducibility has been described as the major obstacle in integrative microarray studies. Discrepancies have arisen due to several variables, including handling procedures, microarray platforms, the number of patients and controls, the body sites sampled, the sample type, the sample procurement methods, the RNA extraction reagent, the analysis bioinformatics software program, and the statistical assessment used.²⁸

A literature review compared the lists of reported publicly available dysregulated genes in keloids and it came up with 25.²⁹ These genes had to have been reported in at least two studies. These 25 genes were either downregulated or upregulated. Another study, in the same time frame, looked at both keloids and hypertrophic scars and came up with 24 dysregulated genes for keloids.²⁸ An additional 40 dysregulated genes were validated using other methods.²⁸

A comparison of these studies highlighted inconsistencies in that only 15 keloid dysregulated genes were common to both. The common genes were

ACAN (aggrecan),

ANXA1 (Annexin A-1),

COL1A1 (collagen type I alpha 1),

COL5A2 (collagen type V alpha 2),

FAP (fibroblast activation protein),

FN1 (fibronectin 1),

IGF2 (insulin-like growth factor 2),

IGFBP7 (insulin-like growth factor binding protein 7),

JAG1 (Jagged1),

OGN (osteoglycin),

SERPINH1 (serpin peptidase inhibitor, clade H1), VCAN (versican),

EGFR, SERPINF1 (serpin peptidase inhibitor, clade F1) and

C5ORF13 (Chromosome 5 open reading frame 13)

As shown by the lack of consensus, even comparing similar studies, makes it very challenging to accurately assign the significance of a putative candidate gene with regard to association with keloid scarring. Importantly, the identified candidate genes by microarray studies thought to be associated with keloid scarring have not been functionally evaluated and as such causative roles for these genes in the development of keloid scarring remain undiscovered and unreported.²⁸

Epigenetics in keloids

An understanding of the role of epigenetics in the pathogenesis of keloid scarring is emerging. This interest in epigenetics has arisen since genetic and environmental factors alone cannot fully explain keloid scarring initiation and progression.

Epigenetics is the study of heritable changes in gene expression, without alteration in DNA sequence. Three main characterized epigenetic modifications are DNA methylation, histone modification, and noncoding RNA-based mechanisms. Figure 2 shows these epigenetic biomarkers in keloid scarring and their inhibitors. Epigenetics is an important mechanism accounting for the pathoclinical complexity observed not only in keloids but also in other skin fibrosing conditions such as systemic sclerosis, heart, lung, and kidney organ fibrosis.³⁰

Figure 2.

Epigenetic biomarkers and their inhibitors in keloid scarring are shown. Histone modification occurs by acetylation or methylation, which then alters the activity of the DNA wrapped around them. HDAC2 deacetylates histones and is overexpressed in keloid tissue, while TSA inhibits histone modification. DNA methylation sites on certain DNA bases repress gene activity mostly by the hypermethylation of promoter regions for the genes CDC2L1, SFRP1, HOXA9, and HOXA10.DNMT methylates DNA, which, in turn, can be inhibited by 5-AZA. Noncoding RNAs (miRNA, lncRNA, and circRNA) exert their effect in keloid scarring by impairing translation into protein or cause mRNA degradation. 5-AZA, 5 aza 2′-deoxycytidine; CDC2L1, cell division cycle2L1; circRNA, circular RNA; DNMT, DNA methyltransferases; HDAC, histone deacetylase; miRNA, microRNA; TSA, trichostatin A. The figures were created with BioRender.com Color images are available online.

DNA methylation

A Genome-Wide Scan for methylation profiles in six keloid tissues highlighted differentially methylated regions with a predominance of hypomethylated genomic landscapes, favoring nonpromoter regions.³¹ Further evidence of differentially methylated CpG sites in fibroblasts from keloid scars was shown in comparison to normal skin and normotrophic scars, thus suggesting that DNA methylation plays an important role in keloid pathogenesis.³²

DNA methylation inhibitors, for example, 5 aza 2′-deoxycytidine, which is also used in the treatment of myelodysplastic syndrome, has been demonstrated to show increased expression of cyclin-dependant kinase (CDK)11p58 protein. In addition, the rate of keloid fibroblast apoptosis was associated with DNA hypermethylation of the cell division cycle2L1 (CDC2L1) gene promoter region.³³

Secreted frizzled-related protein 1 (SFRP1) is a tumor suppressor gene, and it is downregulated in keloid tissue due to hypermethylation of its promoter region. Knockdown of DNA methyltransferase1 (DNMT1) expression demonstrated an upregulation of SFRP1 protein in keloid fibroblasts.³⁴ SFRP1 negatively regulates WNT signaling, which is responsible for keloid cell proliferation by inhibiting the apoptosis of keloid cells.³⁴ This further suggests that SFRP1 might be a candidate therapeutic target for keloid treatment.

Histone modification

DNA is complexed with histone proteins in repeating units called nucleosomes. Histones are affected by post-translational modifications such as lysine acetylation, methylation, phosphorylation, ubiquitination, and sumoylation.³⁵ Furthermore, there are a family of enzymes that mediate these histone modifications such as the histone acetyltransferases, which catalyze the addition of an acetyl group from a donor acetyl-CoA and histone deacetylases (HDACs) which remove the acetyl groups from the histone tails.

A study on keloids implicated HDACs in scarring as HDAC2 was found to be upregulated in both normal and keloid scars.³⁶ In mice models in the same study, HDAC2 was significantly overexpressed in both normal and keloid scar tissue.

Trichostatin A, a known HDAC inhibitor, induced apoptosis in proliferating keloid fibroblasts and simultaneously reduced TGFβ1-induced collagen production, prompting the authors to suggest HDAC inhibitors as potential epigenetic treatment options for keloids.³⁷

This is limited evidence, and more research needs to be done to get more information on the potential role of HDAC inhibitors, but based on the in vivo evidence of the reduction of keloid fibroblast proliferation and the induction of apoptosis, this is a potential treatment avenue that needs to be evaluated.

Noncoding RNAs

Noncoding RNAs are classified according to the length of nucleotides, for example, long noncoding RNA (lncRNA), microRNA (miRNA), circular RNA (circRNA) small nuclear RNA, and small nucleolar RNA. The main known function of noncoding RNA is RNA silencing. Other functions include transcriptional activation, translational regulation, chromatin modification, nuclear-cytoplasmic trafficking, DNA methylation, cell differentiation, and cell cycle regulation. It is only recently that noncoding RNAs have been appreciated as having a role to play in keloid development (Table 4).

Table 4.

Differentially expressed noncoding RNAs in keloids

Noncoding RNA	↑	↓	Reference(s)
miRNA	21	203
	4269	205
	382	2006	³⁸
	199a-5p	205-5p
	21-5p
	214-5p
	424-5p		³⁹
		200c	⁴¹
lncRNA	H19		^42,43
	HOXA11-AS		⁴⁴
		LINC01116	⁴⁶
circRNA	0057452
	0007482
	0020792
	0057342
	0043688		⁴⁷

circRNA, circular RNA; HOXA, homeobox A cluster; LINCO, long intergenic nonprotein coding RNA; lncRNA, long noncoding RNA; miRNA, microRNA.

miRNAs have been identified to be differentially expressed in keloid tissue compared to normal skin tissue. One study discovered 32 differentially expressed miRNA in 12 pairs of keloid/normal tissue,³⁸ while another discovered 264 differentially expressed miRNAs in 3 pairs of keloid/normal tissue.³⁹ Of the quantitative reverse transcription-polymerase chain reaction-verified miRNAs discovered in the first study, miRNA 21 and miRNA200 were upregulated, while miRNA 203 was downregulated. Bioinformatics analyses showed that these miRNAs were associated with TGFβ, p53, and mitogen-activated protein kinase (MAPK) signaling pathways, and of note is that miRNA21 is an oncogenic gene.³⁸

The second study had only miRNA21 in common with the first one, which was also upregulated, while miR199 and miR214 were differentially expressed.³⁹ Other studies have also shown various miRNAs being differentially expressed in keloid tissue in comparison to normal skin.^40,41 Some of these miRNAs, for example, miR200c, have been implicated in certain cancers as well. Expression profiling will lead to a better understanding of the functional roles and the specific pathways affected in keloid etiology, although more research still needs to be done.

Dysregulated expression of lncRNAs is essential during the processes of several human fibrotic diseases, including keloids.⁴² lncRNA H19 is associated with proliferation in cancers and was shown to be overexpressed in eight keloid tissues in comparison to eight normal scars and eight control tissue.⁴² It showed increased proliferative activity of keloid fibroblasts, which may be mediated by mTOR and vascular endothelial growth factor (VEGF).⁴²

Several lncRNA/miRNA axes have been correlated with fibroblast activity. For instance, the H19/miR- 29 axis was studied and it showed that H19 was overexpressed in 80 keloid tissues in comparison to 63 normal scars and 91 controls, while miR29 was underexpressed.⁴³ Silencing of H19 led to overexpression of miR-29 and reduced fibroblast activity through an unidentified mechanism in the COL1A1 pathway.³⁷

In a study in China in 2020, lncRNA homeobox A cluster (HOXA)11-AS was shown to accelerate the progression of keloid formation by the miR-124-3p/TGFβR1 axis by inhibiting cell apoptosis and promoting angiogenesis.⁴⁴ This was concluded after it was shown that HOXA11-AS and TGFβR1 were significantly upregulated while miR-124-3p was downregulated in keloid tissue/fibroblasts in comparison to normal skin tissue/fibroblasts.

HOXA11-AS was also shown in a separate study to aggravate keloid progression through a different pathway, the HOXA11-AS-miR-205-5p-FOXM1 pathway.⁴⁵ Downregulation of the lncRNA LINC01116 inhibited the progression of keloid formation by regulating the miR-203/SMAD5 axis, which might provide a novel target for keloid therapy.⁴⁶ The lncRNA groups, together with its axis, are aberrantly expressed in keloid compared with normal skin tissue, indicating that differentially expressed lncRNAs may orchestrate keloid formation and maintenance.

The role of circRNAs remains largely unknown, but certain circRNAs are significantly upregulated in keloid tissue, that is, hsa_circ 0057452, hsa_circ 0007482, hsa_circ 0020792, hsa_circ 0057342, and hsa_circ 0043688, which were also found to interact with miRNA 29a, miRNA 23a–5p, and miRNA1976.⁴⁷

Therapeutics

There is a plethora of treatment modalities available for the management of keloids, but few remain without recurrence and complications. Common treatment options in keloids are intralesional injection of corticosteroids, use of cryotherapy, excision, radiation, laser, and light-based energy targeted therapies used singly or in combination with other treatment modalities. All these treatment options are insufficient or have undesirable side effects and neither do they target a specific molecule or signaling pathway.

Keloid scarring normally occurs slowly, suggesting therapy may be required for an extended period. It is difficult to consistently measure therapeutic response as well as classify the severity and therefore guide therapeutic strategy. Due to the complexity of signaling pathways in keloid scarring, it seems unlikely that a single therapeutic agent will lead to disease treatment. It may be more beneficial to use combination therapies.

There are emerging genetic and epigenetic therapies in keloid scarring based on advancements from other fields, especially cancers. These therapies are at different stages of development and ultimate approval for human use. These studies mainly stem from the acknowledged benign, yet pseudo-neoplastic cancer-like characteristics of keloids.

Gene therapy with the recombinant adenovirus-mediated double suicide gene cytosine deaminase-thymidine kinase (CDglyTK) has been widely used in cancer. A study demonstrated that the lethal and bystander effects of CDglyTK were marked in keloid fibroblasts.⁴⁸ This supported the notion that recombinant adenovirus-mediated CDglyTK double suicide gene therapy is effective in destroying keloid fibroblasts and therefore may provide a sound scientific rationale for keloid trials in vivo. ⁴⁸ Adenovirus-relaxin gene therapy on keloids has also been shown to attenuate collagen synthesis and MMP expression in keloid fibroblasts, also making it a potential therapeutic agent.⁴⁹

The field of epigenetic therapeutics in hematological cancers is more advanced compared to keloid scarring. Of note is the use of Food and Drug Administration-approved epigenetic biomarker treatment modalities for some cancers. Examples are azacitidine, a DNMT blocker, used to treat myelodysplastic syndrome, and vorinostat, an HDAC blocker used for the treatment of cutaneous T cell lymphoma. Azacitidine has been shown to block DNMT in fibroblasts,³³ suggesting a potential role for epigenetic therapy in keloid scarring.

Studies on the knockdown of certain noncoding RNAs, for example, miR-196a⁴⁰ and lncRNA-ATB,⁴¹ in keloids have demonstrated that these noncoding RNAs and their signaling axis may be a target for therapeutics and may be useful for translational therapeutic interventions.

The use of tyrosine kinase inhibitors, for example, imatinib and nintedanib in pulmonary fibrosis, opened a wide field of innovations and possibilities in targeted treatments for different forms of fibrosis. A controlled study to observe the interaction of sorafenib with keloid fibroblasts in vitro showed it was an effective inhibitor of TGF-β/SMAD and MAPK/extracellular signal-regulated kinase (ERK).⁵⁰ This gave credence to the idea of blocking signaling pathways with analogs and blockers for a desired therapeutic outcome. This is certainly a novel approach for therapeutic targeting of signaling pathways involved in keloid scarring.

Summary

The genetic and epigenetic mechanisms responsible for keloid scarring have not yet been fully elucidated. The therapeutic potential of genetic and epigenetic biomarkers has already been realized in hematological as well solid organ malignancies, which are not amenable to surgical resection, where there are a few epigenetic blockers approved for clinical use. With regard to keloids, evidence is not yet at a comparable level; nevertheless, some of the emerging new technologically enhanced therapeutic approaches appear promising.

These advances will likely enable us to better understand the fibrosis signaling pathways. In the future, further investigations of the interplay between genetics and epigenetics will be most beneficial to elucidate the complex molecular crosstalk and heterogeneity present in keloid pathogenesis.

Take Home Message

Genetics and environmental factors alone have for long been inadequate to explain the onset and development of keloid scarring.

There is currently interest in better understanding the epigenetic mechanisms affecting keloid scarring, mainly DNA methylation, histone modification, and the role of noncoding RNA's.

Early advances in understanding the epigenetic mechanisms affecting keloid scarring show promises of therapeutic potential, even though further research still needs to be carried out.

Keloid scarring depends on an interplay of genetic, environmental, and epigenetic mechanisms.

Footnotes

Acknowledgments and Funding

Professor Bayat is funded by the South African MRC through the Wound and Keloid Scarring (WAKS) Translational Research Unit.

Author Disclosure and Ghostwriting

No competing financial interests exist. The content of the article was expressly written by the authors listed. No ghostwriters were used to write this article.

About the Authors

Dennias Tonderai Nyika, MBChB, FCDerm, MMed (Derm), is a clinical PhD student, interested in the epigenetics of keloids at the Division of Dermatology, University of Cape Town. Nonhlanhla Khumalo, MBChB, PhD, FCDerm, is a Professor and Head of the Division of Dermatology, University of Cape Town. Ardeshir Bayat, BSc (Hons), MBBS, PhD, is Professor and director of the MRC-SA Wound Healing Unit, Hair and Skin Research Laboratory, Division of Dermatology, Department of Medicine, Faculty of Health Sciences and Groote Schuur Hospital, University of Cape Town, South Africa.

Abbreviations and Acronyms

References

Marneros

, Norris

JEC

, Olsen

, Reichenberger

. Clinical genetics of familial keloids. Arch Dermatol, 2001; 137:1429–1434.

Robles

DTMDP

, Berg

DMD

. Abnormal wound healing: keloids. Clin Dermatol, 2007; 2526–32.

Rockey

, Bell

, Hill

. Fibrosis—a common pathway to organ injury and failure. N Engl J Med, 2015; 372:1138–1149.

Igota

, Tosa

, Murakami

, Egawa

, Shimizu

, Hyakusoku

, et al. Identification and characterization of Wnt signaling pathway in keloid pathogenesis. Int J Med Sci, 2013; 10:344–354.

Bettinger

, Yager

, Diegelmann

, Cohen

. The effect of TGF-beta on keloid fibroblast proliferation and collagen synthesis. Plast Reconstr Surg, 1996; 98:827–833.

Brissett

, Sherris

. Scar contractures, hypertrophic scars, and keloids. Facial Plast Surg, 2001; 17:263–272.

Alhady

SMA

, Sivanantharajah

. Keloids in various races: a review of 175 cases. Plast Reconstr Surg, 1969; 44:564–566.

Jfri

, Alajmi

. Spontaneous keloids: a literature review. Dermatology, 2018; 234:127–130.

Yan

, Gao

, Chen

, Song

, Liu

. [Preliminary linkage analysis and mapping of keloid susceptibility locus in a Chinese pedigree]. Zhonghua Zheng Xing Wai Ke Za Zhi, 2007; 23:32–35.

10.

Chen

, Gao

, Liu

, Yan

, Song

. [Linkage analysis of keloid susceptibility loci on chromosome 7p11 in a Chinese pedigree]. Nan Fang Yi Ke Da Xue Xue Bao, 2006; 26:623–625, 37.

11.

Marneros

, Norris

JEC

, Watanabe

, Reichenberger

, Olsen

. Genome scans provide evidence for keloid susceptibility loci on chromosomes 2q23 and 7p11. J Invest Dermatol, 2004; 122:1126–1132.

12.

Santos-Cortez

RLP

, Hu

, Sun

, Benahmed-Miniuk

, Tao

, Kanaujiya

, et al. Identification of ASAH1 as a susceptibility gene for familial keloids. Eur J Hum Genet, 2017; 25:1155–1161.

13.

, Chen

D-M

, Cheng

A-G

, Bao

W-H

, Tang

, Bi

H-S

. Immunogenetic regulation of HLA- II type and CD1a positive cells in peripheral blood of patients with abnormal scar. Chin J Clin Rehabil, 2005; 9:92–94.

14.

W-S

, Cai

L-Q

, Wang

Z-X

, Li

, Wang

J-F

, Xiao

F-L

, et al. Association of HLA class I alleles with keloids in Chinese Han individuals. Hum Immunol, 2010; 71:418–422.

15.

Ashcroft

, Syed

, Arscott

, Bayat

. Assessment of the influence of HLA class I and class II loci on the prevalence of keloid disease in Jamaican Afro-Caribbeans. Tissue Antigens, 2011; 78:390–396.

16.

, Zhang

, Li

, Wang

, Zuo

, Cai

, et al. Association of HLA-DRB1 alleles with keloids in Chinese Han individuals. Tissue Antigens, 2010; 76:276–281.

17.

Brown

, Ollier

WER

, Thomson

, Bayat

. Positive association of HLA-DRB115 with keloid disease in Caucasians. Int J Immunogenet, 2008; 35:303–307.

18.

Brown

, Ollier

, Arscott

, Bayat

. Association of HLA-DRB1* and keloid disease in an Afro-Caribbean population. Clin Exp Dermatol, 2010; 35:305–310.

19.

W-S

, Wang

J-F

, Yang

, Xiao

F-L

, Quan

, Cheng

, et al. Association of HLA-DQA1 and DQB1 alleles with keloids in Chinese Hans. J Dermatol Sci, 2008; 52:108–117.

20.

Nakamura

, Zembutsu

, Takahashi

, Tsunoda

, Nakashima

, Kamatani

, et al. A genome-wide association study identifies four susceptibility loci for keloid in the Japanese population. Nat Genet, 2010; 42:768–771.

21.

Zhao

, Liu

S-L

, Xie

, Ding

M-Q

, Lu

M-Z

, Zhang

L-F

, et al. NEDD4 single nucleotide polymorphism rs2271289 is associated with keloids in Chinese Han population. Am J Transl Res, 2016; 8:544–555.

22.

Ogawa

, Watanabe

, Than Naing

, Sasaki

, Fujita

, Akaishi

, et al. Associations between Keloid Severity and Single-Nucleotide Polymorphisms: importance of rs8032158 as a Biomarker of Keloid Severity. J Invest Dermatol, 2014; 134:2041–2043.

23.

Marneros

. A role for the E3 ubiquitin ligase NEDD4 in keloid pathogenesis. J Invest Dermatol, 2019; 139:279–280.

24.

Zhu

, Wu

, Li

, Wang

, Tang

, Liu

, et al. Association study confirmed susceptibility loci with keloid in the Chinese Han population. PLoS One, 2013; 8:e62377-e.

25.

Velez Edwards

, Tsosie

, Williams

, Edwards

, Russell

. Admixture mapping identifies a locus at 15q21.2–22.3 associated with keloid formation in African Americans. Hum Genet, 2014; 133:1513–1523.

26.

Bayat

, Walter

, Bock

, Mrowietz

, Ollier

WER

, Ferguson

MWJ

. Genetic susceptibility to keloid disease: mutation screening of the TGFbeta3 gene. Br J Plast Surg, 2005; 58:914–921.

27.

Brown

, Ollier

, Arscott

, Ke

, Lamb

, Day

, et al. Genetic susceptibility to Keloid scarring: SMAD gene SNP frequencies in Afro-Caribbeans. Exp Dermatol, 2008; 17:610–613.

28.

Huang

, Nie

, Qin

, Li

, Zhao

. A snapshot of gene expression signatures generated using microarray datasets associated with excessive scarring. Am J Dermatopathol, 2013; 35:64–73.

29.

Shih

, Bayat

. Genetics of keloid scarring. Arch Dermatol Res, 2010; 302:319–339.

30.

O'Reilly

. Epigenetics in fibrosis. Mol Aspects Med, 2017; 54:89–102.

31.

Jones

, Young

, Divine

, Datta

, Chen

, Ozog

, et al. Genome-wide scan for methylation profiles in keloids. Dis Markers, 2015; 2015:943176.

32.

Alghamdi

, Wallace

, Melton

, Moses

, Stevenson

, Al-Eitan

, et al. Identification of differentially methylated CpG sites in fibroblasts from keloid scars. Biomedicines, 2020; 8:181.

33.

Zhang

, Guan

, Chen

, Qian

, Liang

. DNA methylation of the CDC2L1 gene promoter region decreases the expression of the CDK11p58 protein and reduces apoptosis in keloid fibroblasts. Arch Dermatol Res, 2018; 310:107–115.

34.

Liu

, Zhu

, Wang

, Li

, Han

, Liu

, et al. Methylation of secreted frizzled-related protein 1 (SFRP1) promoter downregulates Wnt/beta-catenin activity in keloids. J Mol Histol, 2018; 49:185–193.

35.

Fischle

. Molecular mechanisms of histone modification function. Biochim Biophys Acta, 2014; 1839:621–622.

36.

Fitzgerald O'Connor

, Badshah

, Addae

, Kundasamy

, Thanabalasingam

, Abioye

, et al. Histone deacetylase 2 is upregulated in normal and keloid scars. J Invest Dermatol, 2012; 132:1293–1296.

37.

Diao

, Xia

, Yi

, Wang

, Li

, Xia

, et al. Trichostatin A inhibits collagen synthesis and induces apoptosis in keloid fibroblasts. Arch Dermatol Res, 2011; 303:573–580.

38.

Liu

, Yang

, Xiao

, Zhang

. miRNA Expression profiles in keloid tissue and corresponding normal skin tissue. Aesthetic Plast Surg, 2012; 36:193–201.

39.

Zhong

, Bian

, Lyu

, Jin

, Liu

, Lyu

, et al. Identification and integrated analysis of microRNA expression profiles in keloid. J Cosmet Dermatol, 2018; 17:917–924.

40.

Kashiyama

, Mitsutake

, Matsuse

, Ogi

, Saenko

, Ujifuku

, et al. miR-196a downregulation increases the expression of type I and III collagens in keloid fibroblasts. J Invest Dermatol, 2012; 132:1597–1604.

41.

Zhu

, Bai

, Li

, Zheng

, Guan

, Liu

, et al. Knockdown of lncRNA-ATB suppresses autocrine secretion of TGF-β2 by targeting ZNF217 via miR-200c in keloid fibroblasts. Sci Rep, 2016; 6:24728.

42.

Zhang

, Liu

, Wan

, Peng

, Li

, Qiu

. Long non-coding RNA H19 promotes the proliferation of fibroblasts in keloid scarring. Oncol Lett, 2016; 12:2835–2839.

43.

Wang

, Feng

, Song

, Qi

, Huang

, Wang

. lncRNA-H19/miR-29a axis affected the viability and apoptosis of keloid fibroblasts through acting upon COL1A1 signaling. J Cell Biochem, 2020; 121:4364–4376.

44.

Jin

, Jia

, Luo

, Zhai

. Long non-coding RNA HOXA11-AS accelerates the progression of keloid formation via miR-124-3p/TGFbetaR1 axis. Cell Cycle, 2020; 19:218–232.

45.

, Ma

, Wang

, Gao

. LncRNA HOXA11-AS Aggravates Keloid Progression by the Regulation of HOXA11-AS-miR-205-5p-FOXM1 Pathway. J Surg Res, 2021; 259:284–295.

46.

Yuan

, Sun

, Yu

. Long non-coding RNA LINC01116 accelerates the progression of keloid formation by regulating miR-203/SMAD5 axis. Burns, 2021; 47:665–675.

47.

Shi

, Yao

, Chen

, Yang

, Qian

, Han

, et al. The integrative regulatory network of circRNA and microRNA in keloid scarring. Mol Biol Rep, 2020; 47:201–209.

48.

, Liu

, Zhu

, Yang

, Zhao

, Wang

, et al. Efficacy of recombinant adenovirus-mediated double suicide gene therapy in human keloid fibroblasts. Clin Exp Dermatol, 2008; 33:322–328.

49.

Lee

, Cheon

, Rah

, Yun

. Adenovirus-relaxin gene therapy on keloid; it attenuates collagen synthesis and MMP expression on keloid fibroblast. Burns, 2009; 35:S25.

50.

Wang

, Qu

, Xu

, Wu

, Gao

, Gu

, et al. Sorafenib exerts an anti-keloid activity by antagonizing TGF-β/Smad and MAPK/ERK signaling pathways. J Mol Med, 2016; 94:1181–1194.