Epigenetic modulation of long noncoding RNA H19 in oral squamous cell carcinoma-A narrative review

Abstract

Oral squamous cell carcinoma (OSCC) showed a seemingly increasing incidence in the last decade. In India, despite the use of tobacco decreased rapidly, in the past five years, the incidence pattern of OSCC over gender and age showed a drastic shift. About 51 % of the head and neck cancers are not associated with habits. Studies exploring various contributing factors in the incidence of this malignancy have documented. Recently, the epigenetic factors associated with the induction and progression of OSCC were explored. More than 90 % of the human genome is made up of non-coding transcriptome, which believed to be noises. However, these non-coding RNAs were identified to be the major epigenetic modulators, which raises concern over incidence of carcinoma in non-habit patients. H19 is a long non coding RNA which proved to be an effective biomarker in various carcinoma. Its role in oral squamous cell cancer was not investigated in depth. This review discusses in detail the various epigenetic role of H19 in inducing oral carcinogenesis.

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Epigenetic modulation of long noncoding RNA H19 in oral squamous cell carcinoma-
A narrative review
Peramaiyan Rajendran, Ramya Sekar, Basem M. Abdallah, Shazia Fathima JH,
Enas M. Ali, Selvaraj Jayaraman, Salaheldin Abdelraouf Abdelsalam, Vishnupriya
Veeraraghavan
PII: S2468-0540(24)00020-9
DOI: https://doi.org/10.1016/j.ncrna.2024.01.020
Reference: NCRNA 282
To appear in: Non-Coding RNA Research
Received Date: 29 October 2023
Revised Date: 16 January 2024
Accepted Date: 30 January 2024
Please cite this article as: P. Rajendran, R. Sekar, B.M. Abdallah, S. Fathima JH, E.M. Ali, S.
Jayaraman, S.A. Abdelsalam, V. Veeraraghavan, Epigenetic modulation of long noncoding RNA H19
in oral squamous cell carcinoma-A narrative review, Non-Coding RNA Research (2024), doi: https://
doi.org/10.1016/j.ncrna.2024.01.020.
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Epigenetic modulation of long noncoding RNA H19 in Oral squamous cell carcinoma-A
Narrative review
Peramaiyan Rajendran 1,3#*, Ramya Sekar2,3#*, Basem M. Abdallah1, Shazia Fathima JH3,4; Enas M.
Ali1,5, Selvaraj Jayaraman3; Salaheldin Abdelraouf Abdelsalam1,6 and Vishnupriya Veeraraghavan3
1Department of Biological Sciences, College of Science, King Faisal University, Al-Ahsa 31982,
Saudi Arabia; prajendran@kfu.edu.sa; babdallallah@kfu.edu.sa; eabdelkader@kfu.edu.sa;
sabdelraouf@kfu.edu.sa
2Department of Oral Pathology & Microbiology, Meenakshi Ammal Dental College & Hospitals,
MAHER, Alapakkam Main Road, Maduravoyal, Chennai-95, TN, India;
drramya.oralpathology@madch.edu.in
3COMManD, Department of Biochemistry, Saveetha Dental College & Hospitals, Saveetha Institute
of Medical and Technical Sciences, Velappanchavadi, Chennai-600 077, Tamil Nadu, India;
shaziafathimasyed@yahoo.in; selvarajj.sdc@saveetha.com; vishnupriya@saveetha.com
4Department of Oral Pathology and Microbiology, Ragas Dental College and Hospitals Chennai-
600119, Tamil Nadu, India
5Department of Botany and Microbiology, Faculty of Science, Cairo University, Cairo 12613, Egypt
6Department of Zoology, Faculty of Science, Assiut University, Assiut 71515, Egypt
*Correspondence:
prajendran@kfu.edu.sa; drramya.oralpathology@madch.edu.in;
Tel.: (+966 135899543; (P.R; R.S))
# Equal contribution
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Graphical Abstract
Long non-coding RNA H19 interacting in every hallmark of cancer thereby forming a network of communication
with number of molecules genetically and epigenetically. This indicates its ubiquitous role in oral carcinogenesis
and its importance as a therapeutic target.
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Abstract
Oral squamous cell carcinoma (OSCC) showed a seemingly increasing incidence in the last decade. In India,
despite the use of tobacco decreased rapidly, in the past five years, the incidence pattern of OSCC over gender
and age showed a drastic shift. About 51% of the head and neck cancers are not associated with habits. Studies
exploring various contributing factors in the incidence of this malignancy have documented. Recently, the
epigenetic factors associated with the induction and progression of OSCC were explored. More than 90% of the
human genome is made up of non-coding transcriptome, which believed to be noises. However, these non-coding
RNAs were identified to be the major epigenetic modulators, which raises concern over incidence of carcinoma
in non-habit patients. H19 is a long non coding RNA which proved to be an effective biomarker in various
carcinoma. Its role in oral squamous cell cancer was not investigated in depth. This review discusses in detail the
various epigenetic role of H19 in inducing oral carcinogenesis.
Keywords: H19; lncRNA; microRNA; Biomarker; Oral cancer
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Highlights
• H19 plays a significant role in epithelial mesenchymal transition and in various other pathways that lead
to increased invasiveness.
• H19 is considered an onco-fetal lncRNA, as the product of this gene is RNA, they are abundantly
expressed prenatally.
• H19 can modulate the enzymes, biomolecules and oncogenes and regulate the pathways associated with
glucose metabolism.
• Exosomal H19 play an important role in inculcating stemness in cancer cells by acting as a ceRNA.
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1. Introduction
Oral cancer is the sixth most common cancer in the world [1]. Oral squamous cell carcinoma (OSCC)
is the most common sub-type of malignancy occurring in oral cavity. Studies have stated that OSCC
constitutes about 90% of all oral cancers [2]. They may occur commonly in tongue, buccal mucosa,
gingiva and/or floor of the mouth [3]. Most of the malignant lesions of the oral cavity are due to usage
of tobacco. Although diagnostic and therapeutic field have shown immense development, survival rates
for OSCC are still not satisfactory [4]. Oral cavity is the most accessible region of the body, hence the
potentially malignant changes can be easily appreciated at the initial stages. Early diagnosis of the pre-
malignancies such as oral sub-mucous fibrosis, leucoplakia, erythroplakia and proliferative verrucous
leukoplakia will help in decrease of mortality rates. Global survey shows that about 350,000 new cases
recorded every year [5]. Moreover, the incident rate increased slowly in accordance to the demographic
and clinical data. Heavy smoker’s, and males above 60 years of age are more susceptible for OSCC
development, there is a paradigm shift in incidence of the disease and its pattern resulting in incidence
of the disease among individuals of middle age and without habit [6]. Genetic and environmental factors
play a major role in OSCC incidence and development. Apart from genetics, role of epigenetics in the
initiation and progression of oral cancer was found to be massive; this narrative review briefs the role
of long non-coding RNA H19 in development of OSCC [7].
Human genome consists of about 98% of non-coding transcriptome. These non-coding regions between
the exons or antisense regions gives rise to various types of RNAs [8]. Based on their origin, function
and size of those non-coding RNAs named as microRNA, PIWI-interacting RNAs (P-element Induced
WImpy testis in Drosophila), small nucleolar RNAs, long non coding RNA, ribosomal RNA, transfer
RNA and transcribed ultraconserved regions [9]. The long non-coding RNAs (lncRNA) are the
transcriptome with > 200 nucleotides, which have a revolutionary role in development, diagnosis and
prognosis of oral cancer [10]. They act as a promoter, insulator, enhancer and silencer in various
genomic activity. These lncRNAs are very much time and tissue specific [11]. Various types of long
non-coding RNA in association with OSCC were discovered. TUG 1 (taurine-upregulated gene 1),
HOTTIP (HOXA transcript at the distal tip) and H-19 lncRNAs are associated with epithelial
mesenchymal transition [12]. UCA-1 (urothelial cancer associated 1), NEAT-1(nuclear enriched
abundant transcript 1), LINC RNA ROR (Long intergenic non-protein coding RNA, regulator of
reprogramming), MALAT 1(Metastasis-Associated Lung Adenocarcinoma Transcript 1) showed to be
associated with apoptosis [13]. H-19 is a competing endogenous RNA (ceRNA) that modulates
biological processes such as epithelial-mesenchymal transition and apoptosis [14]. Thus, the role of H-
19 as an epigenetic regulator in oral cancer reviewed in this article.
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2. Classification
LncRNA classified based upon criteria’s such as molecular weight, location, regulation, stability,
biological processes the. As discussed, earlier lncRNAs are transcripts of <200nt (nucleotide). Very
long intergenic non-coding RNAs (vlincRNAs) are those of length 50KB to 1MB. Unspliced sense
PINs- partially intronic RNAs which emerged from transcriptional forests. Totally intronic RNAs-TINs
make about 70% of total non-coding nuclear-encoded RNA and 40-50% of cellular RNAs. Few
transcripts, which regulate the parent gene expression, is referred to as circular intronic ncRNAs
(ciRNAs) and those circular RNA from exons are referred to as exonic circular RNAs (ecircRNAs).
Sense ncRNA, natural antisense ncRNA, mirror antisense, standalone ncRNAs made from 3’UTRs and
transcription start site-associated RNAs are other lncRNAs that are classified based on association with
protein coding genes. Based upon function, non-coding RNAs named as enhancer, upstream antisense,
PROMoter uPstream Transcript, Telomeric repeat-containing RNA and promoter lncRNA. These
ncRNAs involve in transcriptional regulation, chromatin signalling plasticity and dynamics of nuclear
architecture. Based on their resemblance to mRNA, they referred to as long intervening ncRNAs
(lincRNAs) and mRNA like noncoding RNAs mincRNAs [15-23] (Table 1).
H19 is a conserved gene cluster is located at 11p15.5, whose expression is monoallelic. It is an imprinted
gene expressed from maternal allele, that helps in early stage of embryogenesis and growth
development. They also regulate other genes from imprinted gene network like Insulin like growth
factor 2(Igf2), paternally expressed gene 1(Peg1), Slc38a4(solute carrier family 38 member 4). Other
factors such as C/EBP, CTCF insulation, imprinted control region and matrix attachment regions
regulate H19 expression [24]. The primary transcript of H19 gene is processed in a similar way as a
mRNA. H19 gene produces, a 2.4 kb mature transcript. LncRNA H19 transcribed in nucleus and
transported to cytoplasm [25]. They remain untranslated. They are not identified with any open reading
frames; therefore, no protein coding function was identified physiologically. H19 and Igf2 are co-
expressed during embryogenesis in mesoderm and endoderm derived tissues. Myo H or H19 expression
is associated with skeletal and cardiac muscle differentiation [26]. Studies showed that H19 could be
an oncogene/tumour suppressor gene. Recent study has hypothesized that H19 could have a role in cell
cycle, as p53 modulates the expression of H19. It sponges miR138, miR200. Studies have shown that
on silencing H19, increased expression of miR-138 was observed thereby progression and invasion of
OSCC was inhibited [27].
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Table1: Classification of long noncoding RNA based upon various parameters.
Types
About
Example
Reference
Transcript length
Long non-coding RNA
lncRNA
>200 nt
ANRIL,H19, HOTAIR, HOTTIP,
lincRNA-p21, XIST
15
Very long intergenic non-
coding RNAs
50kb to 1MB
Vlinc 21, vlinc_185
15
Annotated protein-coding genes
Intronic ncRNA
TIN,PIN
ANRASSF1
18
Circular ncRNAs
ciRNA
ci-ankrd52, ci-sirt7
19
Sense ncRNA
Sense strand having
exons of protein coding
genes.
senZfp536
16
Antisense ncRNA
asRNA,NAT
SOX21-AS1
16
Functional association with DNA
Enhancer
eRNA
LEENE
17
Promoter
PALR
17
Upstream antisense RNA
uaRNA
17
PROMoter uPstream
Transcript
PROMPT
17
Telomeric repeat-
containing RNA
TERRA
17
Genomic Location and context
Intergenic lncRNAs
lincRNA
20
Intronic lncRNAs
Formed from introns of
the protein coding
genes.
20
Sense and antisense
lncRNAs
NAT, asRNA
aHIF
20
Resemblance to mRNA
mRNA-like noncoding
RNAs
mlncRNA
HOTAIR
21
Long intergenic non-
coding RNA
lincRNA
ANRIL,H19, HOTAIR,HOTTIP
21
Target mechanism of action
Signal
Cell specific expression
Xist
22
Decoy
Binds and evaluate
target protein
PANDA
22
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Guide
Localization of RNP
complex
HOTAIR
22
Scaffold
Helps in localizing
multiple proteins to
configure RNP
complex
HOTAIR,7SL
22
Mechanism of Function
Transcriptional
Regulation
Gene transcription via
transcriptional
interference
H19,MEG3, HOTAIR,ANRIL
23
Post-transcriptional
regulation
Splicing Regulation,
Translational control
H19, PTENP1b, MIAT, MALAT1
23
Other function
Protein localization,
telomere replication,
RNA interference
meiRNA, TERC
23
3. Biogenesis and Degradation
LncRNAs have their biogenesis similar to mRNA. These are conserved, less evolutionary and contained
fewer exons in comparison to mRNA. They are expressed less abundantly possibly due to the repressed
histone modification in their gene promoters. Their 5’ end has m7G caps and the 3’ end has
polyadenylated tails. Many studies stated that formation, processing and degradation of lncRNAs are
linked to their fate and function [28]. Most of the lncRNAs are inefficiently processed by
phosphorylation dysregulated polymerase II (Pol II) and held back in nucleus rest are spliced and
transported to cytoplasm by nuclear RNA export factor 1(NXF1). Those poorly transcribed RNAs in
nucleus are degraded by exosomes (nuclear). LncRNA with U1snRNA (U1 small nuclear RNA) binding
motif recruit consecutive nucleoprotein (U1RNP) and establish association with polymerase resulting
in number of non-coding RNA formation [29]. Factors in cis and trans such as U1 small nuclear
ribonucleoprotein (U1snRNP) and heterogenous nuclear ribonucleoprotein K (hnRNPK) also enhance
nuclear localization. Due to shorter PPT (polypyrimidine tract) few lncRNAs undergo inefficient
splicing. Other RNPs such as PPIE (peptidylprolyl isomerase E) downregulates the splicing process and
contribute in nuclear localisation. Increased accumulation of lncRNA occurs in chromatin when SPT6,
a pol II associated elongation factor is altered functionally [30]. Loss of SPT6 will eventually result in
active transcription of lncRNA genes instead of protein coding genes, subsequently causes increased
lncRNA accumulation to result in DNA damage associated R-loops. LncRNA in cytoplasm showed to
interact diversely with many RNA binding proteins (RBP) and some with ribosomes. Pseudo 5’
untranslated regions help in association of lncRNA with ribosomes [31]. Those lncRNA showed to have
half reduced half-life period. Many lncRNAs mobilised into mitochondria, without known function.
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Exosomes also identified with few lncRNAs, RNA array studies reveal that 29% of lncRNA were
unstable with half-life period of 2h or even less. Cytoplasmic lncRNAs are more stable than lncRNAs
of nucleus. LncRNAs structurally mimic mRNAs and might degrade in the same way as mRNAs by
processes such as deadenylation, decapping and exo/endonucleolytic degradation [32]. Studies
hypothesized, that lncRNA decay by nonsense-mediated mRNA decay (NMD), STAU1-mediated
mRNA decay (SMD), no-go mRNA decay (NGD) and non-stop mRNA decay (NSD). Decay
machineries recruited by miRNAs and RNA binding proteins (RBPs) are also thought to be involved in
lncRNA decay. Evidences suggested that H19 sequences are targeted by let-7. Overexpression of let-
7a was reported to reduce H19 levels. However, the exact mechanism of degradation of lncRNA yet to
be explored [33].
3.1Biogenesis and Degradation of H19:
H19 is transcribed from the H19 gene, which is maternally imprinted and paternally expressed. It is
transcribed by RNA polymerase II. The expression of H19 and its reciprocally imprinted gene, insulin-
like growth factor 2 (IGF2), is regulated by an imprinting control region. Methylation status at this
region affects the expression of both genes. Post-transcriptional modifications and processing, like 5'
capping, 3' polyadenylation, and splicing, prepare the nascent H19 RNA for export from the nucleus to
the cytoplasm [28].
The stability of H19 RNA is regulated by various factors, including RNA-binding proteins, microRNAs
(miRNAs), and other regulatory molecules that influence its half-life. H19 can act as a molecular sponge
for miRNAs, regulating their availability to target mRNAs. Some miRNAs can target H19 for
degradation, affecting its stability. Endonucleases and exonucleases can degrade RNA molecules,
including lncRNAs like H19. Enzymes like RNase P and RNase R are involved in RNA degradation
pathways. Various cellular signals and stress conditions can influence the degradation rates of RNAs,
including H19, by altering the activity of degradation machinery or affecting RNA-protein interactions.
The intricate regulation of H19's biogenesis and degradation contribute to its diverse functions in
cellular processes. Understanding these processes is crucial for unravelling the roles of H19 in
development, diseases, and potential therapeutic interventions [34].
4. Expression and Function of H19
In situ hybridisation technique evaluation of extraembryonic tissue of mouse shows that expression of
H19 was evident by E5.5 and in embryo soon after implantation by E6.5- biallelical [35]. Expression
of H19 identified throughout the foetal development in comparable levels of β-actin expression. Igf2
expressed in a similar pattern of H19. Their expression reduced in all tissues except skeletal muscles
after birth [36].
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In-vivo study of mouse models helps in identification of regulatory elements of the expression pattern
of imprinted genes. Imprinting control region (ICR), differentially methylated domain (DMD),
differentially methylated region (DMR) identified in H19 transcription start site between -4 and -2 kb.
Enhancers and promoters also controlled the H19 gene by [37].
The expression pattern of the long non-coding RNA H19 is dynamic and tissue-specific, playing
essential roles during development and in various physiological processes. Few key aspects of its
expression pattern are
Developmental Expression:
H19 is highly expressed in the placenta and during embryonic development, particularly in
various embryonic tissues, including muscle, liver, and placenta. Its expression pattern is
drastically reduced after birth and altered during chronic diseases [35]. H19 is subject to parental
imprinting, meaning that its expression is allele-specific and depends on whether it is inherited
from the mother or the father. In most tissues, the maternally inherited H19 allele is active, while
the paternally inherited allele is silenced due to DNA methylation at the imprinting control region
(ICR) [37].
Tissue-Specific Expression:
H19 is highly expressed in the placenta during development, where it plays a role in regulating
placental growth and function. H19 involves in muscle development and differentiation,
particularly in regulating myogenesis. H19 found to be expressed in the liver, where it may
contribute to liver development and function[38].
Disease and Pathological Conditions:
Altered expression levels of H19 was observed in various cancers such as breast cancer,
hepatocellular cancer. In some cancers, H19 is overexpressed and contributes to oncogenesis by
acting as an oncogene, regulating cell proliferation, migration, invasion, and apoptosis. While in
retinoblastoma it acts as a tumour suppressor gene with low expression pattern. H19 implicated
in diabetes-related complications and metabolic disorders affecting insulin signalling and
pancreatic beta-cell function [39].
Regulation:
H19 expression is regulated by epigenetic mechanisms, including DNA methylation at the
Imprinting Control Region-ICR, histone modifications, and chromatin remodelling. Various
transcription factors and signalling pathways can modulate the expression of H19 in response to
developmental cues or environmental stimuli. The tissue-specific and developmentally regulated
expression pattern of H19 underscores its diverse roles in normal physiology and disease. Its
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aberrant expression in certain pathological conditions highlights its potential as a biomarker or
therapeutic target in various diseases, including cancer and metabolic disorders[40].
H19 helps in limiting cell proliferation and body weight. H19 was found to be involved in number of
cancers such as hepatocellular carcinoma, bladder carcinoma, choriocarcinomas, breast cancer, oral
cancer and in association with syndrome such as Beckwith wideman syndrome [41]. They showed to
have upstream effect in hormonal regulation and downstream effect in metabolism, tissue invasion,
migration and angiogenesis. H19 levels were down regulated in type 2 diabetes [42]. H19 via TGF-β
pathway induces osteoarthritis, and rheumatic arthritis. H19 was also observed to be a mediator in stress
induced inflammation. Increased expression of H19 was observed in hypertrophied cardiomyocytes of
mouse and in cardiac ischemia. Conversely, in ischemic stroke. H19 modulates Notch1 and inhibit the
transcriptional activity of p53, hence increasing the susceptibility of the disease [43]. Many animals
model experimental studies conducted conclude that deletion of a 13-kb and 3-kb region resulted in
overgrowth. H19∆13 mutants cause deletion of DMR thereby expression of Igf2 allele is enhanced.
H19 expression was hyper-pronounced during development. Although no viable data has been collected
on its role in embryogenesis/development. Studies done so far conclude that H19 could play a major
role in development. H19 also functions as competing endogenous RNA (ceRNA), encoding proteins
necessary for myotube formation [44].
Deletion mutation in H19 results in overgrowth phenotype, this could be due to modification of
chromatin structure as a result of deletion. In addition, this results in change in accessibility of Igf2
promoter, it could be also due to interference of H19 RNA in loss of Igf2 imprinting. H19 functions as
both in cis and in trans [45]. Trans function of H19, it was evident in H19∆13 and H19∆3 mutants,
change in amount of methylation of Igf2 DMRs was evident in opposite allele thus suggesting of trans
effect. H19 also modulate polysomes thereby affecting the translation of Igf2. Many in-vitro
experiments were done to show the trans effect of H19. Study by Hansen et al showed that H19 could
interfere with Igf2 regulation and translation [46].
5. H19 and Epigenetics
The expression of H19 depends upon the DNA methylation, addition of CH3-methyl group to the
cytosine in a CpG dinucleotide of the residual DNA. CpGs occurs in islands and enzyme DNA methyl
transferases process DNA methylation. Methyl transferases can create new methylation patterns [47].
Conventionally, genes with methylated promoter cannot be expressed. Other epigenetic modifications
apart from methylation are demethylation, acetylation and deacetylation of specific amino acid residues
[48]. Paternal chromosome has methylated H19 promoter thence no transcription takes place. Enhancers
are short DNA sequences, which helps in transcription of genes [49]. Animal studies on deletion of
enhancers showed decrease in H19 expression especially in endoderm derived tissues. Chromosome
11p15 has two imprint control regions ICR1, ICR2. ICR1 corresponds to differentially methylated
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region-H19 DMD which has abundant CpG residues. H19 ICR1 plays a vital role in transcription of
H19 as well as preventing enhancers from accessing Igf2 promoters in maternal chromosome. Studies
done on immune-precipitated cells suggested that paternal H19 allele was hypoacetylated in comparison
to maternal allele which could affect H19 imprint [50]. Mir-let-7 seem to be in association with H19 in
the process of inhibition of insulin receptor and lipoprotein lipase [51]. H19 encodes mir-675 that might
affect the expression of TGF-β. H19 also downregulates hypertrophy factor via mir-675 in
cardiomyocyte hypertrophy [52]. They also inhibit autophagy of cardiomyocyte by promoting
phosphorylation of mammalian target of rapamycin. H19 showed to regulate mir-200, mir-138 in
association with epithelial mesenchymal transition in hepatocellular/colorectal cancers [53] (Figure 1)
Figure 1
Figure 1: Various epigenetic modulation of H19 in influencing the initiation and progression of
tumorigenesis in oral squamous cell carcinoma. Image created using biorender.com (accessed on 18
May 2023). H19, H4K20Me3 (histone H4 lysine 20 tri-methylation), and H4K16Ac (histone H4 lysine
16 acetylation) are all related to epigenetics and chromatin regulation, but they play distinct roles in
gene expression and chromatin structure. H19, H4K20Me3, and H4K16Ac, they all contribute to the
complex regulatory mechanisms that control gene expression and chromatin structure in cells.
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6. H19 and other systemic diseases
Role of H19 associated with many systemic diseases. H19 expression strongly correlated with many
aging processes, inflammation, diabetes, pre-eclampsia, osteoarthritis, Rheumatoid arthritis,
Alzheimer’s, cardiovascular diseases, obesity, auto immune diseases, liver diseases, and chronic kidney
disease. They also modulate telomere length, heterochromatin formation, senescence, proteostasis, cell
cycle regulation, stem cell differentiation and intracellular communication [54-56].
The diverse involvement of H19 in these diseases and biological processes highlights its multifaceted
roles in health and disease, making it a promising target for understanding disease mechanisms and
potential therapeutic interventions.
Disease Associations
H19's expression is linked to cellular senescence and aging-related mechanisms. It plays roles in
modulating telomere length, which is associated with cellular aging, and it contributes to regulating
senescence pathways [57]. Inflammatory processes often involve dysregulation of H19 expression,
contributing to the regulation of inflammatory responses and cytokine production. Dysregulation of
H19 expression has been observed in pre-eclampsia, a pregnancy complication characterized by high
blood pressure and signs of damage to other organ systems. H19 plays roles in joint health and
inflammation, contributing to the pathogenesis of arthritis conditions [58]. Studies have suggested
associations between H19 dysregulation and Alzheimer's disease progression, though the exact
mechanisms are still being explored [59]. Long non-coding RNAs (lncRNAs) play a pivotal role in
governing cellular biological processes via diverse molecular mechanisms. Their prevalence within the
cardiovascular system emphasizes their substantial role in cardiovascular functions and diseases. One
such lncRNA, H19, exhibits high evolutionary conservation and is notably abundant in cardiac and
vascular tissues. This abundance underscores its crucial contribution to preserving the balance and
proper functioning of the cardiovascular system [60]. H19 expression is linked to adipogenesis and
adipocyte function, influencing obesity-related pathways. Its involvement in modulating immune
responses can contribute to autoimmune disease pathogenesis. H19 dysregulation showed to be
associated with liver diseases such as hepatocellular carcinoma and fibrosis, as well as chronic kidney
disease progression [55].
Biological Processes
The role of H19 in modulating telomere length and senescence pathways links it to cellular aging and
longevity. H19 influences proteostasis mechanisms, contributing to protein homeostasis and cellular
function. H19 participates in cell cycle regulation, impacting cell proliferation and differentiation
processes. It has been implicated in regulating stem cell differentiation in various tissues. H19's
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regulatory functions extend to modulating intracellular communication pathways, affecting cellular
signalling and responses [57].
7. Role of H19 in Cancer
H19 is a lncRNA that has been studied extensively in various contexts, including cancer. Here are some
potential reasons, why H19 might be of interest in comparison to other ncRNAs in OSCC [61].
Imprinted Gene Regulation: The imprinted nature of H19 adds another layer of complexity to its role
in cancer. As an imprinted gene, H19 exhibits mono-allelic expression, meaning only one copy inherited
from either the mother or father is actively expressed, while the other copy is silenced. This exclusive
expression pattern is critical in normal development and growth regulation. However, in cancer,
disruptions in this imprinting can occur, leading to abnormal expression levels or loss of imprinting.
Such alterations can contribute to dysregulated cellular behaviours, including uncontrolled
proliferation, evasion of growth suppressors, and resistance to cell death—fundamental traits of cancer
cells. H19's involvement in these imprinting abnormalities positions it as a compelling candidate for
understanding the underlying mechanisms driving cancer initiation and progression. Its disrupted
imprinting might serve as an early indicator or contribute to the cellular changes that lead to malignancy.
Moreover, investigating the regulatory mechanisms behind H19 imprinting alterations could provide
valuable insights into potential therapeutic targets or strategies aimed at restoring normal imprinting
patterns, potentially impacting cancer progression and treatment outcomes. [62].
Epigenetic Regulation: The link between H19 and epigenetic modifications, particularly in chromatin
remodelling, is a crucial aspect of its involvement in cancer biology. Epigenetic alterations, which
include modifications like DNA methylation, histone modifications, and chromatin remodelling, are
pivotal in regulating gene expression patterns without changing the underlying DNA sequence. H19's
association with epigenetic modifications, such as DNA methylation changes in its regulatory regions
or its influence on chromatin structure, highlights its role in modulating the expression of various genes
involved in cancer-related pathways. Understanding how H19 contributes to these epigenetic alterations
can provide significant insights into cancer development and progression. For instance, aberrant
methylation patterns around the H19 locus or its interaction with chromatin-modifying enzymes might
influence the expression of tumor-suppressor genes or oncogenes, impacting cancer cell behaviour.
Furthermore, therapies targeting epigenetic modifications have gained attention in cancer treatment.
Investigating the relationship between H19 and these modifications could unveil potential avenues for
therapeutic intervention. By elucidating the mechanisms by which H19 influences epigenetic changes,
researchers may identify new targets for precision medicine approaches aimed at restoring normal
epigenetic regulation in cancer cells. [63].
Interactions with microRNAs: H19's role as a molecular sponge or a competing endogenous RNA
(ceRNA) is a fascinating aspect of its function in regulating gene expression networks. MicroRNAs
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(miRNAs) are small non-coding RNAs that typically bind to the 3' untranslated regions (UTRs) of target
mRNAs, leading to their degradation or inhibition of translation. H19, as a long non-coding RNA,
contains miRNA binding sites, allowing it to competitively bind to miRNAs, essentially acting as a
sponge that sequesters these miRNAs. By doing so, H19 can modulate the availability and activity of
miRNAs, preventing them from binding to and regulating their target mRNAs. This competitive binding
mechanism could have significant downstream effects on gene expression networks. If H19 sponges
certain miRNAs, it leads to an increase in the levels of miRNA targets that would have otherwise
suppress by these miRNAs. Consequently, this can affect various cellular processes by altering the
expression of genes involved in pathways related to proliferation, apoptosis, differentiation, and other
critical functions. In the context of cancer, dysregulation of this ceRNA network involving H19 and
miRNAs can contribute to the aberrant expression of genes associated with oncogenesis or tumour
suppression. Understanding these interactions provides insights into the intricate regulatory
mechanisms governing cancer development and progression, offering potential therapeutic targets for
manipulating these networks to modulate cancer cell behaviour [64] (Figure 2).
Metastasis and Invasion: H19's precise role in cancer is complex and multifaceted, several studies have
suggested its involvement in promoting cancer metastasis and invasion. The mechanisms underlying
this promotion of metastasis and invasion by H19 are diverse. The ability of H19 to influence various
cellular processes, including proliferation, migration, and epithelial-mesenchymal transition (EMT),
contributes to its impact on cancer progression. By regulating downstream target genes through its
interactions with miRNAs or other molecular pathways, H19 can affect the expression of genes
involved in metastasis and invasion. It might modulate factors that control cell adhesion, motility, and
the ability of cancer cells to invade surrounding tissues and migrate to distant sites. Additionally, H19's
involvement in signalling pathways associated with metastasis, such as those involving TGF-beta or
Wnt signalling, further supports its role in promoting these aggressive cancer phenotypes [61].
Understanding the precise mechanisms through which H19 contributes to metastasis and invasion is
crucial for developing targeted therapies aimed at disrupting these processes. H19 being a part of
complex regulatory networks involving other molecules known to be important in OSCC, that might be
chosen for study due to its potential to provide a comprehensive view of these networks.
The process of carcinogenesis is highly regulated by inflammation, many factors such as tumour
necrosis factor-α and interleukin 6 (IL-6) play a remarkable role in carcinogenesis. H19 is considered
to be an oncofetal lncRNA, as the end product of this gene is RNA, they are abundantly expressed
prenatally as well decreased expression was observed post-nataly [61] . They are hyper expressed in
cancer cells, post-nataly. H19 was first reported to be upregulated in bladder cancer and was identified
to be the predictive marker in early diagnosis of recurrence [65]. In esophageal cancer and colorectal
cancer, the H19 levels were upregulated, while in breast and lung cancer it is down regulated. It has
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been found to be associated with cholangiocarcinoma, H19 sponges mir-let-7 and induce IL-6 to initiate
carcinogenesis [66]. In breast cancer H19 sponges with mir-152 and promotes proliferation in cancer
cells. H19 modulates the expression of p53 thereby affecting cell repair as well apoptosis. H19
associated with epithelial mesenchymal transition (EMT), as well mesenchymal epithelial transition
[67]. H19 acts as a microRNA sponge and results in EMT in colorectal cancer [68]. H19 has been
shownshowed to inhibit E-cadherin subsequently promoting EMT. Role of H19 in number of pathways
that promotes tumorigenesis was identified, such as p38/MAPK, NF-κB, PI3-AKT/mTOR. cMyc, an
oncogene was identified to regulate H19. Many studies performed to relate H19 in every stage of cancer
progression. Thereby, it seems to be a potent marker in diagnosis and prognosis [69-71].
Oncogenic and Tumor Suppressor Activities: The dual nature of H19 as both an oncogene and a tumor
suppressor gene in different types of cancers adds complexity to its role in cancer biology. Its diverse
functions are context-dependent, varying based on the cellular environment, specific cancer type, and
the intricate signalling pathways involved. For example, H19 exhibits oncogenic properties by
promoting cancer cell proliferation, invasion, and metastasis through various mechanisms like acting
as a ceRNA, modulating gene expression, or influencing critical signalling pathways. On the other hand,
in certain contexts or cancer types, H19 demonstrated tumor-suppressive effects by inhibiting cell
proliferation, inducing apoptosis, or regulating pathways associated with suppressing tumor growth and
progression. This context-dependent behaviour of H19 emphasizes the importance of considering its
multifaceted roles in different cancers. Factors such as the microenvironment, genetic background, and
interactions with other molecules within the cellular network can influence whether H19 acts as an
oncogene or a tumor suppressor [61,62].
Understanding the specific conditions under which H19 exerts its oncogenic or tumor-suppressive
functions is crucial for targeted therapeutic interventions. It highlights the need for personalized
approaches in cancer treatment, taking into account the specific molecular characteristics of the tumor
and its microenvironment. The intricate nature of H19's roles in cancer underscores its potential as a
diagnostic biomarker and therapeutic target. Further research aimed at elucidating these context-specific
roles will be essential for harnessing the full potential of H19 in cancer management. [72,73].
Diagnostic and Prognostic Biomarker Potential: H19 is an intriguing biomarker with diagnostic and
prognostic potential in certain cancers. As a long non-coding RNA, it plays an important role in various
cellular processes, including cancer development and progression. Studies have shown its aberrant
expression in cancers like bladder, breast, colorectal, and gastric cancer, among others. For diagnostics,
detecting elevated levels of H19 in bodily fluids or tissues might indicate cancer presence. It could serve
as a non-invasive screening tool, aiding in early detection when treatment outcomes are generally more
favourable. Regarding prognosis, the expression levels of H19 might correlate with cancer
aggressiveness, metastasis, and patient survival rates. Higher H19 expression could indicate a poorer
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prognosis or treatment response, helping clinicians tailor therapies or anticipate disease progression
[74,75].
However, further research is crucial to validate H19's efficacy, specificity, and sensitivity across various
cancer types and stages. While promising, its clinical application as a reliable biomarker requires
extensive validation and standardization to ensure accuracy in diagnosis and prognosis.
Figure 2
Figure 2: H19 influencing miR675-5p; miR675-3p; miR21; miR200; miR-200 for regulating
transcription, translation and Gene silencing thereby causing various epigenetic triggers in initiation
and promotion of oral cancer along with the genetic factors. Image created using biorender.com
(accessed on 18 May 2023).
8. H19 and oral cancer
H19 has a vital role in regulating the proteins involved in carcinogenesis. It sponges number of miRNAs
and regulate them. Their tissue specific expression suggest that they could act as an early
prognostic/diagnostic marker [76]. Study by Yu et al reported the role of H19 in aberrant activation of
fibroblast thereby playing a crucial role in fibrogenesis- oral sub mucous fibrosis (OSMF) [77].
Overexpression of H19 in OSMF is associated with disease progression by sponging miR-29b. Study
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done by Guan et al showed that the upregulation of H19 was in correlation with poor overall survival
rate and disease-free survival [78]. Kou et al studied the role of H19 as competing endogenous RNA by
sponging miRNA let- 7a in tongue squamous cell carcinoma (TSCC) [79]. Their findings concluded
that H19 plays an important role in regulation of TSCC migration and invasion. Ghapanchi et al [80]
studied H19 gene polymorphism in oral squamous cell carcinoma (OSCC) tissues and analysed H19
rs217727T allele mutation. They concluded significant association between the OSCC susceptibility
and H19 polymorphism. Guo et al also analysed the association between OSCC and H19 gene
polymorphism sites such as rs217727, rs2735971, rs2839698 and rs3024270 [81]. Their research
indicated that rs217727 polymorphism was statistically correlated with OSCC incidence. These results
were in support of Ghapanchi et al study. Zhang et al analysed clinic-pathological characteristics and
expression pattern of H19 and enhancer of zest homolog 2 (EZH2) in TSCC [82]. They concluded that
H19 promotes the progression of TSCC via EZH2. Zhao et al investigated the effect of H19 on
migration and invasion in oral cancer cell line [83]. The regulation of H19, mir-107and cyclin dependent
kinases 6 were analysed. It was concluded that inhibition of H19 decreased migration and invasion.
Vishwakarma et al studied the expression of various lncRNA by real time PCR in OSCC tissues and
determined that H19 was downregulated in OSCC tissues [84]. The expression of H19 was associated
with smoking status. Lee et al studied transcription and methylation status of H19 by PCR in OSCC
cell line and patient tissue [85]. They observed hypomethylation of H19 in OSCC tissue in comparison
to normal mucosa and inferred that it could act as a prognostic biomarker.
Zhou et al revealed, the major role of H19 depletion has a in growth and invasion of malignant cells in
oral cancer [86]. Thereby they could act as a potential target as they bind with ZEB1 protein which aids
in epithelial mesenchymal transition. Yang et al, examined the profiles of H19 in normal fibroblast and
cancer associated fibroblast of OSCC tissues [87]. Knockdown of H19 suppressed MAPK pathway and
miR-675-5p which h involved in glycolysis pathway and concluded that it could act a biomarker in
diagnosis. El-Naggar et al assessed the imprinting status of H19 and IGF2 in head and neck cancer by
real time PCR and suggested their role in tumorigenesis of head and neck malignancy [88]. Hong et al
analysed the aberrantly expressed lncRNA in OSCC [89]. They also quantitatively analysed the
expression levels of H19 and correlated with their clinical characteristics and prognosis.
Downregulation of H19 inhibited proliferation, invasion, migration and epithelial mesenchymal
transition in OSCC cells in-vitro. They also detected correlation between miR-138 and H19. H19 by
competing with miR-138 they regulated the expression of (EZH2) enhancer of zeste homolog 2. Wang
et al studied the expression of lncRNA H19 in tongue squamous cell carcinoma and he inferred that
overexpression of H19 causes reduced proliferation of tumour cells [90]. They explained the role of
H19 through miR-675-5p/GPR55 axis. Esteves et al correlated loss of allele specific expression of IGF2
and H19 with differentially methylated region in head and neck squamous cell carcinoma [91]. They
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detected H19 expression levels were in relation with recurrence indicating that it could act as a potent
prognostic marker (Table 2).
Table 2: Detailed tabulation of works done by various authors in the oral potentially malignant
disorders and oral squamous cell carcinoma in relation to the long non coding RNA H19.
S.No
Study
Inference
Reference
1
Yu et al 2021 on myofibroblasts in osmf
Overexpression of H19 in OSMF is
associated with disease progression
77
2
Guan et al 2016 on HNSCC
upregulation of H19 was in correlation
with poor overall survival rate and
disease-free survival.
78
3
Kou et al 2019 on TSCC
H19 as competing endogenous RNA.
H19 plays an important role in
regulation of TSCC migration and
invasion.
79
4
Ghapanchi et al 2020 on OSCC
They concluded significant association
between the OSCC susceptibility and
H19 polymorphism.
80
5
Guo et al 2017 on OSCC
Their research indicated that rs217727
polymorphism was statistically
correlated with OSCC incidence.
81
6
Zhang et al 2017 on TSCC
H19 promotes the progression of TSCC
via EZH2.
82
7
Zhao et al 2019 on oral cancer cells
It was concluded that inhibition of H19
decreased migration and invasion.
83
8
Vishwakarma et al 2020 on OSCC
Determined that H19 was
downregulated in OSCC tissues.
84
9
Lee et al 2021 on OSCC
They observed hypomethylation of H19
in OSCC tissue in comparison to normal
mucosa and inferred that it could act as
a prognostic biomarker.
85
10
Zhou et al 2020 on Oral cancer cells
Depletion of H19 has a major role in
growth and invasion of malignant cells
in oral cancer.
86
11
Yang et al 2021 on CAF in oral cancer cell
H19 suppressed MAPK pathway and
miR-675-5p which has been involved in
glycolysis pathway and concluded that
it could act a biomarker in diagnosis.
87
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12
El-Naggar et al 1999 on HNSCC
Imprinting status of Igf2 and H19 could
enhance tumorigenesis
88
13
Hong et al 2018 on OSCC
Downregulation of H19 inhibited
proliferation, invasion, migration and
epithelial mesenchymal transition in
OSCC cells in-vitro.
89
14
Wang et al 2021 on TSCC
Overexpression of H19 causes reduced
proliferation of tumour cells.
90
15
Esteves et al 2005 on HNSCC
H19 expression levels were in relation
with recurrence indicating that it could
act as a potent prognostic marker
91
9. H19 and Cancer associated fibroblast
The important component of tumor micro environment, cancer associated fibroblast (CAF) play an
important role in promoting tumorigenesis. Cross talk between the CAF and tumor cells are believed to
cause tumor progression [92]. The role of H19 in promoting glycolysis was identified. H19 showed to
modulate the enzymes, biomolecules and oncogenes and regulate the pathways associated with glucose
metabolism [93]. Inhibitors of enzymes that play a role in glycolysis. The functional role of H19 in
modulating glycolysis in oral CAFs were analysed and it was observed that knockdown of H19 impairs
glucose uptake and as well hinders lactate secretion. Studies have also shown that H19 mediates
glycolysis via miR-675-5p in oral CAFs. CAFs attain their stemness of cancer cells via wnt/βcatenin
signalling pathway, studies showed that exosomal H19 play an important role in transferring the cancer
stem cell property by acting as a ceRNA through miR-141 [94].
10. H19 role in inducing Stemness
Role of H19 in embryogenesis and normal tissue homeostasis are well established. Studies reported that
silencing of H19 in embryonic endothelial precursors results in failure of haematopoietic stem cell
formation. Role of H19 in various tumor progression processes such as genomic stability, stemness,
migration, invasion and chemoresistance were researched. H19 was also found to induce symmetrical
and asymmetrical renewal of cancer stem cells (CSCs) in various cancers. Its role in modulating various
miRNAs such as let-7c thereby inducing stemness in cancer cells are studied. Role of H19 in inducing
CSCs of oral squamous cell carcinoma is one inadequately explored field [93,95].
11. H19 as a Biomarker
Biomarkers are those substances which help in early diagnosis, prognostic indicator and therapeutic
target. These substances could be a protein, mRNA, DNA or miRNAs [96]. H19 acts as a potential
diagnostic and therapeutic marker in diseases such as breast cancer, gastric cancer, colorectal cancer,
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cardiovascular diseases, multiple myeloma, bladder cancer [97]. Authors such as Lee et al, Yang et al
and Esteves et al suggested H19 as a diagnostic and prognostic biomarker in OSCC [85, 87, 91].
Researches was also done to study the efficacy of H19, as a potent therapeutic target to suppress the
invasion and metastasis in oral cancer. More studies associating H19 expression pattern with various
stages of oral cancer and dysplasia has to be done to understand the expression pattern of this non coding
RNA [98]. Studies suggesting the prognostic and therapeutic value of H19 in oral cancer are devoid as
they are associated with the metastatic and recurrence potential of the malignancy. H19 was associated
with the miR let-7 and promotes metastasis, by promoting the activity of HRAS gene [99]. Role of H19
in metastasis via the cancer associated fibroblasts studied in colorectal cancer. Their role in metastasis
of oral squamous cell carcinoma has not been detailed yet [53]. H19 as a potential biomarker in OSCC
has to be studied in more details, as early diagnosis will help in tailor made treatment plan thereby
preventing invasive treatments [100].
12. Conclusion
LncRNAs play a vital role in the progression of OSCC. Their role in metastasis has not been explored
well. Role of H19 as a competing endogenous RNA in various pathways associated with the
tumorigenesis of oral cancer should to be studied in more details. Single nucleotide polymorphism of
H19 is associated with many cancers as well oral cancer. H19 is hence regarded as a potent biomarker
in diagnosis and prognosis of Oral cancer. Many studies correlate oral cancer and H19 expression. This
review provides an overview for the role of H19 in oral cancer. H19, the potential lncRNA can
effectively play a major role in tailoring precision therapeutics in OSCC. Unfortunately, literature
support the expression pattern of this non coding RNA is not sufficient with respect to head and neck
cancer. More research studies are required to be conducted to establish the axis of relationship between
various carcinogenesis signalling pathway and H19. Thus, its potential role as a biomarker and
therapeutic target can be established.
Funding: The author acknowledged the Deanship of Scientific Research, Vice Presidency at King Faisal
University Saudi Arabia for financial support under the annual funding track [Grant 3,444].
Institutional Review Board Statement: Not applicable.
Informed Consent Statement: Not applicable.
Data Availability Statement: Not applicable.
Acknowledgments: Deanship of Scientific Research at King Faisal University, Saudi Arabia.
Conflicts of Interest: The authors declare no conflict of interest.
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Author Contributions:
CONCEPTION: P.R. and R.S.
INTERPRETATION OR ANALYSIS OF DATA: R.S, P.R, S.F.J.H, V.V;
PREPARATION OF THE MANUSCRIPT: P.R, B.M.A, E.M.A S.A.A;
REVISION FOR IMPORTANT INTELLECTUAL CONTENT: P.R, B.M.A
SUPERVISION: P.R
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