Africa Medical College

Department of Pharmacy

A Literature Review on Perspective of Histamine and Its

Receptor

For the Partial Fulfillment as the Requirement of the Bachelor

Degree in Pharmacy (B. Pharm)

By: Tihitina Getachew

Advisor: Teklay G/Mariam

August, 2020
Addis Ababa, Ethiopia

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Acknowledgements

First of all, I would like to thank GOD whose grace made me enable to carry out this review.
Secondly I would like to express my deepest appreciation to all those who provided me the
possibility to complete this paper. A special gratitude I give to my advisor Mr.Teklay whose
contribution in stimulating suggestions and encouragement helped me to coordinate my literature
review especially in writing this review paper.

Contents

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Acknowledgements..........................................................................................................................2
Contents...........................................................................................................................................3
Abbreviations and acronyms...........................................................................................................3
List of tables and figures..................................................................................................................5
Summary..........................................................................................................................................6
1. Introduction..................................................................................................................................7
2. Molecular pharmacology of histamine-4(H4) receptor...............................................................8
2.1. Overview of Histamine Receptors........................................................................................8
2.2. Classification, structure, cellular expression and function of histamine receptors...............8
2.3. Overview of pathophysiological of histamine and histamine receptors.............................10
2.3.1. Over view of physiological role of histamine and its receptors...................................10
2.3.2. Common pathological role of histamine and its receptors...........................................12
2.3.2.1. Histamine-4 receptor (H4R) and inflammatory disorders........................................13
2.3.2.2 H4R and Autoimmune disorders................................................................................15
2.4 H4R gene..............................................................................................................................17
2.5 H4R expression, protein structure and post-translational modifications.............................18
2.6 H4 Receptor signaling..........................................................................................................20
3. Histamine-4 receptor as therapeutic target................................................................................25
3.1. Overview of potential therapeutic targets...........................................................................25
3.2. Histamine receptors as therapeutic target of inflammation................................................26
4. Future perspectives....................................................................................................................28
5. Conclusion and recommendations............................................................................................29
References......................................................................................................................................30

Abbreviations and acronyms

ATP Adenosine triphosphate

CNS Central nervous system

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COX Cyclo oxygenase
DC Dendritic cell
EGFR Epidermal growth factor
ELS Extra cellular loops
GPCR G- Protein Coupled Receptors
HA Histamine
HR Histamine receptor
H1R Histamine-1 Receptor
H2R Histamine-2 Receptor
H3R Histamine-3 Receptor
H4R Histamine-4 Receptor
HDC Histidine Decarboxylase
ILS Intra cellular loops
NK Natural killer
ROS Reactive oxygen species
RA Rheumatic arthritis
RAMH R-a-methyl histamine
SNP Single nucleotide polymorphism
T-rep T - Regulatory
MC Mast Cells

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 List of tables and figures

Tables

Table 1: summary of histamine receptors cellular expression, histamine affinity, associated

GPCRs and cellular effectors which mediate histamine signaling pathways................................24

Figures

Figure 1: Schematic representation of the expression of histamine receptors in inflammatory and

immune cells .................................................................................................................................12
Figure 2: Influence of histamine on mucosal-associated immune cell subsets.............................15
Figure3: Schematic representation of the H4R gene location on chromosome 18 and an overview
of the exon/introns distribution the different splice variants.........................................................18
Figure 4: Snake plot of the human H4R protein............................................................................21
Figure 5: the schematic representation of proposed signaling pathway for the production of IL-13
and RANTES by H4R in human mast cells...................................................................................23
Figure 6: Histamine receptor signal transduction..........................................................................24
Figure 7: Chemical structures of H4R ligands and their binding to other histamine receptor
subtypes….27

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Summary

The short-acting endogenous amine (Histamine (2-[4-imidazolyl]- ethylamine)), which is widely

distributed throughout the body, is synthesized by histidine decarboxylase (HDC), an enzyme
which decarboxylates the semi-essential amino acid L-histidine which is recognized for its
effects in the immediate-type hypersensitivity response(i.e., increased vascular permeability,
smooth muscle contraction, activation of nociceptive nerves, wheal-and-flare reaction, and itch
response), the pathological relevance of increased histamine levels at diseased sites is less well
understood in most of autoimmune disorders.

In this review, the various signaling mechanisms, physiological involvement and pharmacologic
approaches targeting histamine-4 receptors in inflammatory and autoimmune disorders were
included in this review work. Furthermore, there are a number of published papers on this areas
especially before 2011after H4R development in 2001 but the field is still very energetic,
reflected by the numerous publications focusing on distinct signaling mechanisms,
pathophysiological involvement and pharmacologic approaches targeting histamine-4 receptors
in inflammatory and autoimmune disorders showing promising role of H4R on those disorders
and development of new drugs for the upcoming human clinical trials.

Keywords: histamine, H4R, inflammation, ligands, signaling

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1. Introduction

Histamine pharmacology has experienced a new beginning over the last few years, and this
short-acting endogenous amine (Histamine (2-[4-imidazolyl]- ethylamine)), which is widely
distributed throughout the body, is synthesized by histidine decarboxylase (HDC), an enzyme
which decarboxylase the semi-essential amino acid L-histidine was originally discovered at the
beginning of the 20th century, and first chemically synthesized by Windaus and Vogt in 1907.
As soon as its discovery the first biological functions of histamine were reported by Dale and
Laidlaw in 1910 and it is found ubiquitously in the mammalian organism and it acts in various
physiological and pathophysiological processes as chemical mediator and neurotransmitter in
peripheral and central tissues. Mast cells and basophile are important sources of HA, which is
released from granule stores in response to several triggers recognized as an important
immunosuppressant and immunoregulatory mediator in cell-mediated inflammatory disorders as
well the histamine receptor subtypes offered a great research field to medicinal chemists of
numerous generations [1].

The histamine H4 receptor (H4R) is the youngest member of the histamine receptor family.
Based on its predominant expression pattern in hematopoietic cells, the H4R is considered to be
an interesting drug target for inflammatory disorders such as allergy and asthma. Differences
between H4R orthologs in combination with available three-dimensional G protein coupled
receptor (GPCR) models have guided site-directed mutagenesis studies to gain insight in ligand
binding and receptor activation. In addition, ongoing characterization of H4R-mediated signaling
in transfected and native cells contributes to further unravel the (patho-) physiological functions
of H4Rs [2]. The number of potential therapeutic targets which have applications in the H4R
field reached in between 2012 and 2018 most of these compounds belong to the already
described 2-amino-pyrimidine and the indole classes suggesting that H4 drug discovery projects
are primarily focusing on these chemo types.

Therefore, this review paper aimed at the various signaling mechanisms, physiological
involvement and pharmacologic approaches targeting histamine-4 receptors in inflammatory and
autoimmune disorders.

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 2. Molecular pharmacology of histamine-4(H4) receptor

2.1. Overview of Histamine Receptors

Histamine receptors (H1R to H4R) are GPCRs, which are a large receptor characterized by an
extracellular N-terminal domain followed by 7 transmembrane alpha-helical domains connected
by 3 intracellular and 3 extracellular domains. G protein refers to a protein that is able to bind
guanyl nucleosides and has the ability to act as an exchanging factor by catalyzing replacement
of the bound guanosine diphosphate by guanosine triphosphate. GPCRs bind diverse ligands,
varying in size from small molecules to large proteins, and include light-sensitive compounds,
pheromones, hormones, and neurotransmitters. Histamine receptors contain seven
transmembrane domains and belong to the rhodopsin-like family of GPCRs. Each HR is
associated with specific G-alpha subunits, which result in distinct molecular signaling cascades
and diverse modes of action following histamine binding four histamine receptors have been
characterized so far (H 1 R, H 2 R, H 3 R and H 4 R), and an important role for histamine has
been identified in several physiological and pathological responses. In particular, histamine
acting through H 1 R or H 2 R plays a primary role in the development of allergic disorders and
in the regulation of gastric acid secretion, respectively. Although all histamine receptors are
expressed in the brain, the preferential expression of H 3 R in the central nervous system
underlies its pivotal role in the regulation of basic homeostatic and higher functions, including
cognition, arousal, circadian and feeding rhythms. Finally, the role of H 4 R in the modulation of
several aspects of the immune response is now increasingly being appreciated [3].

2.2. Classification, structure, cellular expression and function of histamine receptors

Histamine is a ubiquitous chemical messenger that can be released from a variety of cells (e.g.
mast cells, entero chromaffin-like cells, neurons) to act on one of four histamine receptors:-H 1, H2,
H3 and H4. The classification of histamine receptors has resulted from rigorous pharmacological
analysis of the response of different isolated tissues and cells to histamine. At the present time, the
four histamine receptors identified by addition of histamine receptor ligands and by observing
tissue response which are not been added to by the application of more recent cloning techniques.
Indeed, the nucleotide and protein sequences of the histamine H 3 and H4 receptors have only just
been elucidated. In contrast to the other histamine receptors, splice variants of the H 3 and H4

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receptors have been detected. There is some preliminary pharmacological evidence to suggest
heterogeneity within some of the known types of histamine receptor (reviewed by the
subcommittee in reference , but this heterogeneity may arise from species variation , splice
isoforms and/or oligomerization and awaits the development of specific pharmacological tools.
the four histamine receptors can be described as heptahelical G-protein coupled receptors. They
transduce extracellular signals through various G proteins, which function as mediators between
the cell surface receptors and the intracellular second messenger systems. Histamine and its four
receptors (H1R–H4R) represent a complex system of immune regulation with distinct effects
dependent on receptor subtypes and their differential expression [3]. The H1 receptor is widely
distributed throughout the body, with well-documented expression in the CNS, smooth muscle,
sensory nerves, heart, adrenal medulla, and immune, endothelial, and epithelial cells. The H1
receptor mediates most of the postsynaptic effects of histamine within the central nervous system.
Moreover, through its activity at H1 receptors, histamine stimulates smooth muscle contraction in
the respiratory and gastrointestinal tract, stimulates sensory nerves leading to pruritus and
sneezing, and increase vascular permeability leading to edema. Simultaneous activation of H1 and
H2 receptor can also result in hypotension, tachycardia, flushing, and headache. The H2 receptor
is also widely expressed and can be found in gastric mucosal cells, heart, CNS, immune cells, and
smooth muscles of the airway, vasculature, and uterus. H2 receptor activation stimulates
hydrochloric acid secretion from the acid secreting parietal cells of the gastric mucosa, leads to
smooth muscle relaxation in the vasculature and airways, increases cardiac rate and contractility
and mediates some of the immune modulatory effects of histamine [4].

The histamine H3 receptor is found mainly in the central nervous system (basal ganglia,
hippocampus and cortical areas), but can also be found in the peripheral nervous system, airways,
the cardiovascular system, and the gastrointestinal tract. Acting through presynaptic H3 receptor,
histamine regulates its own release as well as the release of other neurotransmitters such as
noradrenalin, dopamine, serotonin, acetylcholine, and gamma amino-butyric acid. In the lower
airways, H3 receptors are located on postganglionic cholinergic nerves and defend against excess
broncho constriction and in the upper airways, histamine may play a role in nasal congestion
through its activity at H3 receptors. The H4 receptor has been detected in bone marrow, peripheral
blood, spleen, thymus, lung, gastrointestinal tract, liver, peripheral nerves, and central neurons.
Nevertheless, cells that clearly express functional H4 receptors are mainly hematopoietic and

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 include: mast cells, eosinophils, basophiles, dendritic cells, and T cells. H4 receptor activation
induces calcium mobilization in mast cells and mediates mast cells migration towards histamine.
Moreover the receptor plays a significant role in regulating dendritic and T-cell function.
Generally H1R and H2R showed wide expression throughout the gastrointestinal tract and the
expression pattern of H4R was mostly similar [4],which is a recently described histamine
receptor, with selective expression pattern restricted to medullary, peripheral hematopoietic cells
and keratinocytes it is assumed to play an important pro-inflammatory role in various disorders
[5-6].

2.3. Overview of pathophysiological of histamine and histamine receptors

2.3.1. Over view of physiological role of histamine and its receptors

Histamine is one of the most extensively studied biological amines in medicine. It stimulates smooth
muscle contraction and gastric acid secretion, increases vascular permeability, functions as a
neurotransmitter, and plays various roles in immune modulation, allergy, inflammation,
haematopoiesis and cell proliferation. Histamine is involved in the regulation of many physiological
functions including cell proliferation and differentiation, hematopoiesis, embryonic development,
regeneration, and wound healing. Within central nervous system it affects cognition and memory, the
regulation of sleep cycle, energy and endocrine homeostasis (figure 1).

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Figure 1: Schematic representation of the expression of histamine receptors in
inflammatory and immune cells [7]
Histamine secreted from various cells following challenge by pathogens, modulates the function
of neutrophils and other phagocytes mainly through the activation of H1R and H2R. One of the
most important microbicidal tools of phagocytes is the production of reactive oxygen species
(ROS) and Some groups confirmed that histamine inhibits ROS production by isolated
neutrophils or macrophages through H2R [8]. In addition to this histamine induces the
proliferation and migration of airway smooth muscle cells and epithelial cells, which are key
characteristics of airway remodeling of asthma, through EGFR ligand (amphiregulin and HB-
EGF) release [9]. Histamine (HA) is also considered as a neurotransmitter in the adult
mammalian CNS, where it regulates via both pre- and post-synaptic mechanisms a variety of
central responses and functions, such as wakefulness, feeding, drinking, the neuroendocrine
system, body temperature, analgesia, motor activity and has been reported to have both

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anxiolytic and anxiogenic effects as well as involved in embryonic development, considered as
one of the first neurotransmitters to be present in the CNS, reaching its maximum level at early
embryonic development to steadily decrease until birth [10]. Furthermore histamine promotes the
release of ATP from human subcutaneous fibroblasts via Panx1 hemi channels, leading to
calcium mobilization from internal stores and cell growth through the cooperation of H1 and
P2Y receptors (most probably of the P2Y1 subtype) activation. Targeting the pathways leading
to nucleotide release and the purinergic cascade, consisting in metabolizing E-NTPDase and P2
purinoceptor activation, may be useful in designing novel therapies toward the modulation of cell
signals between fibroblasts, nociceptors, and inflammatory cells, which underlie the
pathogenesis of painful musculoskeletal diseases with widespread involvement of the
subcutaneous connective tissue, such as fibromyalgia [11].

2.3.2. Common pathological role of histamine and its receptors

Histamine is not only the major mediator of the acute inflammatory and immediate
hypersensitivity responses (i.e., increased vascular permeability, smooth muscle contraction,
activation of nociceptive nerves, wheal-and-flare reaction, and itch response), but has also been
demonstrated to affect chronic inflammation and regulate several essential events in the immune
response. It can influence numerous functions of the cells involved in the regulation of immune
response and hematopoiesis including macrophages, dendritic cells, T lymphocytes, B
lymphocytes and endothelial cells. These cells express histamine receptors and also secrete
histamine, which can selectively recruit the major effectors cells into tissue sites and affect their
maturation, activation, polarization, and effectors functions leading to chronic inflammation for
example histamine up regulates the expression of H1R and H3R in astrocytes clarify the
involvement and mechanism of astrocyte activation in neuroinflammation [12].

Histamine regulates antigen specific Th1 and Th2 cells, as well as related antibody isotype
responses and prevents the expression of late differentiation antigens in keratinocytes and
strongly decreases the expression of tight junction and desmosomal proteins, leading to the
formation of a defective skin barrier [13].

Histamine acting through its receptor (HR) type 2 positively interferes with the peripheral
antigen tolerance induced by T regulatory (T Reg) cells in several pathways. The diverse effects
of histamine on immune regulation are due to differential expression and regulation of 4

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histamine receptors and their distinct intracellular signals. In addition, differences in affinities of
these receptors are highly decisive on the biological effects of histamine and agents that target
histamine receptors. Hisatmine-4 receptor modulate eosinophils migration and selective
recruitment of mast cells leading to amplification of histamine-mediated immune responses and
eventually to chronic inflammation as well as it is involved in dendritic cell activation and T cell
differentiation which documents its immuno-modulatory function. The characterization of the
H4 as the immune system histamine receptor directed growing attention towards its therapeutic
exploitation in inflammatory disorders, such as allergy, asthma, chronic pruritus and autoimmune
diseases. Due to complicated receptor pharmacology, the lack of a reliable antibody and limited
availability of primary cells expressing the receptor the physiology of this receptor is still poorly
understood.

2.3.2.1. Histamine-4 receptor (H4R) and inflammatory disorders

The immune response is governed by stringent effectors and regulatory processes that normally
result in protection from infection and tolerance of offensive environmental antigens. But, in
patients with inflammatory diseases, the activated immune response results in a chronic pro-
inflammatory state characterized by activated innate pathways with aberrant expansion and
polarization of T-helper cells, or regulatory T (T reg) lymphocyte populations so that knowing
appropriate controlling factors that augment protective immune responses while limiting tissue
damage is urgently required of which one factor that influence immune reactivity and receiving
more attention as an immunomodulator is histamine (2-[4-imidazolyl]- ethylamine).

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Figure 2: Influence of histamine on mucosal-associated immune cell subsets

Luminal sources of histamine include the diet and bacteria, while mast cells and basophiles are
the primary sources of tissue-derived histamine. Epithelial cells and immune cells express
histamine receptors. Activation of dendritic cell (DC) H1R supports Th1 lymphocyte
polarisation, while H2R inhibits Th1 and Th2 lymphocyte polarization and supports the
expansion of regulatory lymphocytes [14].

The one which recently identified histamine receptor, H4R appeared to have a selective
expression pattern restricted to medullary and peripheral hematopoietic cells and it is assumed to
play an important pro-inflammatory role in various diseases, including bronchial asthma, atopic
dermatitis, and pruritus [5]. Stimulation of H4R resulted in induction of proliferation of human
keratinocytes, this response to histamine in patients with atopic dermatitis might play an

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important pathophysiological role, which could be alleviated by H4R antagonists [6,15,16],
controls NK cell migration in the skin during early allergic inflammation and inducer of NK cell
activation which leads to CCL17 release from dendritic cells which in turn can attract TH2 T
cells into the site of inflammation [17]. In addition to this histamine4 receptor (H4R) have a
threefold higher affinity thanH1R/H2R for histamine and are found almost mostly on neurons
and immune tissues plays a crucial role in migraine pathogenesis, sustaining the neurogenic
inflammation pathway associated with interaction between mast cells (MC) and calcitonin-gene
related protein (CGRP) results in sensitization of trigeminal afferents and trigeminal ganglia
[18], clinical severity of skin disease like psoriasis also enhanced in patients who have anti-drug
anti-bodies and the level of TNF-alpha increased even though patients are treated with TNF-
alpha inhibitors which leads requirement of monitoring the plasma drug level and the presence of
anti-drug anti-bodies in daily clinical practice [19-21].

Generally histamine 4 receptor (H4R) mediates the histamine-induced migration or cell shape
change of dendritic cells, mast cells and eosinophils and synergizes with other chemotactic
receptors to enhance the migration and cell shape change of eosinophils, play a critical role in a
number of different acute and chronic inflammatory disorders. Consistent with its expression in
the nervous system, the histamine H4 receptor was demonstrated to mediate both histamine and
antigen-specific induced acute responses even though histamine have role in augmenting
inflammatory effect on different cells and tissues as described earlier it is characterized by
exerting multiple opposing or complementary effects on cells with immune, vascular, muscular,
neuronal, epithelial, and endocrine function of which histamine through the H4R exhibits a
crucial role in tumors progression. Therefore, H4R ligands offer a novel therapeutic potential as
adjuvant for breast cancer treatment [5], autocrine activation of H4Rs on the mast cells (MC)
membrane by MC-derived histamine leads to sequentially PKC and ALDH2 activation,
reduction of toxic aldehyde-induced MC degranulation, decreased rennin release, prevention of
RAS activation, reduction of NE release, and ultimately alleviation of reperfusion arrhythmias
[22].

2.3.2.2 H4R and Autoimmune disorders

The immune system evolved to discriminate self from non self but when there is activation of
histamine -4 receptor which is highly expressed in the major effectors of innate immune cells
like complement, granulocytes, monocytes/macrophages, NK cells, mast cells, and basophiles

15
and major effectors of adaptive immune cells like B and T lymphocytes those cells become
highly activated and mediate autoimmunity when those cells are activated disturbances of these
regulatory systems could potentially lead to over suppression of the immune system for example,
resulting in a higher susceptibility to cancer and infectious diseases, or over activation of the
immune system which on its turn may lead to a higher risk for inflammatory or autoimmune
diseases, and there is an imbalance between pro- and anti-inflammatory path-ways has been
proposed to play an important role in the pathogenesis of several neuro developmental disorders
including autism [23].

The involvement of various receptors present on immune cells like B cells, T cells, basophiles
and mast cells associate with lipid rafts on ligand binding and initiate signaling cascades leading
to inflammation. Furthermore, disrupting lipid raft integrity alters lipo polysaccharide-induced
cytokine secretion, e.g. signaling, and B-cell and T-cell activation so that the integrity of rafts
and spatial segregation of receptors in rafts regulates a variety of signal transduction pathways
and specific targeting of lipid rafts could be used as a potential therapeutic target in
inflammatory and autoimmune diseases [24].

In addition to this, inflammatory cytokines and chemokines are believed to be involved in the
development of rheumatic arthritis (RA). Therefore, these cytokines are regarded as targets for
RA treatment. In the arthritic model examined in this study, inflammatory cytokines such as IL-
1β, IL-6 and TNF-α and chemokines such as MCP-1 were shown to contribute to the
development of arthritis and there is up-regulation of H4R expression in diseased tissues has
been postulated to occur in rheumatic arthritis, where by its expression level is attenuated by
histamine-4 receptor antagonists [25]. A role for histamine in malignant cell growth has long
been proposed. The presence of H1, H2 and H4 receptors has been confirmed on cancer cells from
the colon. For example, in Caco-2 cells, histamine induces an up-regulation of cyclooxygenase
(Cox-2) expression, which can be antagonized by H 2 and H4 blockers, Whereas H1 blockers are
intensively applied for only in allergy-induced itch, and not when other mechanisms are
involved. In mouse models of itch, H4 blockers show additive efficacy and combined H1–H4
blockade might be a new therapeutic avenue for the treatment of itch [25].

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2.4 H4R gene

The protein encoded by the human histamine H4R gene (i.e. HRH4) shows very low (~19%)
amino acid sequence homology to the histamine H1 and H2 receptors and shares ~37% identity
with the H3R and its transcription results in a mRNA molecule of 3.7kb. The open reading
frame is 1173bp in size and encodes 390 amino acids. (Figure 3)
The HRH4 contains two introns of 7867 BP and >17500 BP, dividing the actual coding region
into three exons (amino acids 1-65, 66 and 120-390[2].

Figure3: Schematic representation of the H4R gene location on chromosome 18 and an

overview of the exon/introns distribution the different splice variants.
The gene is located on position q11.2 and consists of three exons and two introns. The regions
that encode the typical 7 Trans membrane (TM) domains of the full length H4R (390) are shown
above the sequence. The splice variant H4R (302) does not contain exon-2 and recognizes an
alternative acceptor site (dotted vertical line) in between the TM4 and TM5 coding sequence.
The H4R (67) variant passes over exon-2, continues at the normal acceptor site of exon-3, which
subsequently results in a frame shift and premature stop codon [2].

Molecular cloning of H4R showed a very limited homology with all the known histamine
receptors (approximately 31% with H 3 R and only 23% and 22% with H 1 R and H 2 R,
respectively) [7]. The H4R protein has three splice variants which code the 390, 302- and 67-
amino acid isoforms .Among the two exonic single nucleotide polymorphism (SNPs)
(rs11665084 and rs11662595) the rs11665084 is presents both in the 390- and 302-amino acid

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isoforms, but rs11662595 is present only in the 390-amino-acid-long isoforms. None of these
SNPs are present in the 67-amino-acid-long isoforms. Nevertheless, researches finding were
mostly focused with two SNPs (rs527790 and rs487202) downstream of the HRH4 gene. This is
an intergenic region, the nearest flanking genes of HRH4 are IMPACT (upstream) and
LOC390843 pseudo gene (downstream), however, they do not overlap with HRH4 and are not in
linkage disequilibrium with it. The localization of these SNPs suggests the existence of some
possible functional regulatory elements in this region [26].

2.5 H4R expression, protein structure and post-translational modifications

The histamine H4 receptor (H4R) is the youngest member of the histamine receptor family
which is expressed in a variety of tissues such as bone marrow, spleen, peripheral blood, thymus,
small intestine, colon, heart and lung with predominant expression pattern in hematopoietic cells
and considered to be an interesting drug target for inflammatory disorders such as allergy and
asthma [27]. In human tissues hH4R expression was detected in bone marrow, peripheral blood,
spleen, thymus, small intestine, colon, heart and lung at mRNA levels and these mRNA studies
present discrepancies in the expression pattern in distinct cells, which underline the importance
of anti-H4 receptor antibodies in the immunohisto-chemical detection of the receptor. The
majority of H4R expressing cells indeed belong to the hematopoietic system (neutrophils, mast
cells, eosinophils, basophiles, dendritic cells, monocytes and T cells). Similar to the other
members of its family, the H4R also belongs to the large G-protein coupled receptor (GPCR)
family, consisting of seven transmembrane-spanning helices. And expressed in cells that are key
players in regulation of immune disorders: mast cells, eosinophils, basophiles, dendritic cells,
and several subsets of T cells [28]. H4 R is also highly expressed by Th2 cells or CD4 T cells
stimulated with IL-4. Moreover, polyclonally activated peripheral blood mononuclear cells or
Th2 cells stimulated with H 4 R agonists up regulated IL-31 mRNA, a cytokine involved in skin
allergic inflammation and the induction of pruritus [20]. Additionally, the H4R was found to be
expressed in endocrine cells in the gastrointestinal tract, on dermal fibroblasts and nerves of the
human nasal mucosa. Besides the existence of H4R in the enteric nervous system in rodents,
expression in the CNS was also confirmed [1]. The H4 receptor expressed on dendritic cells,
CD4+ and CD8+ T cells appears to control cytokine and chemokines production [29]. Histamine
receptors belong to the family of GPCRs. These receptor proteins consist of seven cell

18
membrane-spanning alpha-helices that are connected by three extracellular loops (ELs) and three
intracellular loops (ILs). The N-terminal tail is located extracellular and the C-tail intracellular.
The histamine receptors have several conserved structural motifs that are common to the class A
(rhodopsin-like) GPCRs, including the highly conserved residues Asn1.50, Asp2.50, Arg3.50,
Trp4.50, Pro5.50, Pro6.50 and Pro7.50 (43). Like in most other GPCRs, TM3 and the EL2 of
H4R are presumably connected by a disulphide bridge between Cys3.25 and Cys45.50. The
extracellular N-terminal tail contains two asparagines (Asn1.21 and Asn1.25) that are predicted
to be involved in post-translational glycosylation, whereas the intracellular C-tail is possibly
anchored to the cell membrane through palmitoylation of the Cys7.69 residue. In addition, like
most GPCRs, H4R has an additional helix 8 that is located intracellularly and does not span the
cell membrane [2]. The H4 receptor (protein) shows a relatively high level of homology with the
H3 receptor. Comparable to what has been observed for H 3, spliced isoforms occur for H4, and as
is the case for H3, important species variation characterizes the H 4 system; consequently, ligand
activities vary from species to species. As a consequence of the H 3/H4 homology, several H3
ligands show effects on H4 as well (this is especially true for imidazole derivatives); the opposite
also applies [2].

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Figure 4: Snake plot of the human H4R protein.
The cell membrane-associated H4R has 7TM helices and several conserved structural motifs that
are common to class A (rhodopsin-like) GPCRs. A disulphide bridge between the conserved
Cys3.25 and Cys45.50 (gray circles) connects the first EL/TM3 and the second EL. The N-
terminal extracellular tail contains two asparagines (Asn1.21 and Asn1.25) (bold) that are
possibly involved in post-translational glycosylation. The intracellular C-tail is presumably
anchored to the cell membrane through palmitoylation of the Cys7.69 residue (bold). Residues
that have been shown to be important in ligand binding and/or receptor activation as well to be
responsible for observed differences in ligand binding between species are shown in black circles
with white text [2].

2.6 H4 Receptor signaling

The physiological regulatory character in cellular events and immunobiological effects of

histamines are modulated by activation through four distinct G-protein-coupled histamine
receptors designated as H1, H2, H3 and H4) that are differentially expressed in various tissues
and cells [3]. Cells of both the innate and adaptive immune system can be regulated by

20
histamine, binding of histamine to four subtypes of histamine receptors shape and define the
nature of histamine effects in immunological responses. Histamine binding to the H1R leads to
activation of phospholipase C (PLC), which produces 1, 2-diacylglycerol and inositol- 1, 4, 5-
trisphosphate, subsequently resulting in activation of protein kinase C (PKC) and release of
calcium ions from intracellular stores while, H2R is an adenyl cyclase-coupled GPCR, which
stimulates cAMP production and downstream effects are mediated by protein kinase A (PKA)
and the transcription factor cAMP response element-binding protein. Histamine binding to H3R
results in the activation of mitogen-activated protein kinase (MAPK) pathways, protein kinase B
(PKB) and an increase in calcium ion concentration [13].

The H4R activation utilizes different signaling pathways for the production of various
inflammatory mediators ,ERK is a well-characterized mitogen activated protein kinase (MAPK)
and is primarily associated with the regulation ,proliferation, anti-apoptosis, differentiation and
cytokine gene expression [30], histamine receptor, H4R, shares some molecular,
pharmacological and signal transduction features with H3R [31] and triggers calcium release
from internal storage, activates PI3K/Akt pathways, and inhibits cAMP/PKA activation [18,31].

21
Figure 5: the schematic representation of proposed signaling pathway for the production of
IL-13 and RANTES by H4R in human mast cells
The activation of H4R induces the phosphorylation of ERK1/2, Akt and nuclear translocation of
NFjB-p65. Both ERK1/2 and Akt signaling pathways are involved in H4R mediated IL-13
production, whereas, ERK1/2 alone drives RANTES/CCL5 production via H4R [30]. It is
promising that we now start to identify the signaling pathways from the periphery to the central
nervous system since the brain itself is complex organ where several neurotransmitters interact,
such as the noradrenergic and histaminergic systems which reveals the interaction between the
brain and the periphery, in addition to this genes coding H1R and H4R mediate the activation of
MAPKs and CREB in a cooperative manner, thus, signaling pathways relaying on others than on
calcium and cAMP may be involved in this regulation which are not yet identified and require
further investigation.

22
Figure 6: Histamine receptor signal transduction
HRs is GPCRs which bind to heterotrimeric G proteins, consisting of alphabeta- gamma
subunits, following ligand binding. The classical pathway involves the association of specific G
alpha subunits (Galpha, Galphas and Galphai/o) to activate distinct molecular signaling cascades
[32].

23
Table 1: summary of histamine receptors cellular expression, histamine affinity, associated
GPCRs and cellular effectors which mediate histamine signaling pathways [18]
Sub Gene G Effectors Expression Histamine
type protein Affinity
H1R 3p25 Gα q/11 Ca2+, Epithelial,smoothvascular ˜1µg/mL
PKC endothelial,immune,glial,neuronalcells
H2R 5q35.5 Gα5 PKA Epithelial,endothelial,immune,gastric,neuronal ˜3µg/mL
Gαq/11 PKC cells
H3R 20q13.33 Gi/o PKA, Neuronal cells,cardiac synaptosomes,GI tract ˜1ng/mL
NHX
H4R 18q11.2 Gi/o PKA, Immunetissue(lung,spleen,guts,,lymphnode,bone ˜2ng/mL
Ca 2+ marrow,leukocytes)
PKC=Phosphokinase C;PKA=Phosphokinase A;NHX=Sodium-hydrogen exchanger

24
3. Histamine-4 receptor as therapeutic target

3.1. Overview of potential therapeutic targets

Since the discovery of the H4R, numerous ligands have been identified that bind the receptor and
affect downstream signaling pathways of those ligands. Histamine derivatives ( example:-5-
methyl-imbutamine), In dole, benzimidazole and benzothiazole analogs, Amino-pyrimidines
analogs, Quinazoline derivatives, Rotationally constrained aminopyrimidines and radio ligands
are the most commonly used drugs with greater therapeutic application in different inflammatory
and autoimmune disorders. Azines (Furopyrimidines, Pyrazolopyridines, Pyrazolopyrimidines,
Thienopyrimidines, Quinazoline and Cycloheptyl pyrimidines), especially pyrimidines
derivatives, are a promising class of H4R antagonists/inverse agonists and their potential
therapeutic utility of constitutes an attractive target in the search for new drugs which suggests
that H4R antagonists/inverse agonists can be useful for the treatment of allergic rhinitis, asthma,
rheumatoid arthritis, atopic dermatitis, idiopathic chronic urticaria, inflammatory pain,
neuropathic pain or osteoarthritis pain [33]. Several efforts have been made to design and
synthesize H4R selective (inverse) agonists and antagonists. Considering the amino acid
similarity to the H3R, especially in the ligand binding pocket that is formed by the TM domain,
it is not surprising that the majority of imidazole-containing H3R ligands have affinity for the
H4R as well. Examples include R-a-methyl histamine (RAMH), immepipimmetridine and
methimepip. Small changes in ligand structure result in great differences in histamine receptor
subtype specificity [2].

25
Figure 7: Chemical structures of H4R ligands and their binding to other histamine
receptor subtypes [2]

3.2. Histamine receptors as therapeutic target of inflammation

Histamine H4 receptor (H4R) has been shown to be involved in various inflammatory conditions
and the ligands acting on H4R show therapeutic potential in various diseases. Treating patients
with breast cancer using histamine-4 receptor agonists like clozapine or JNJ28610244 produce a
concentration dependent inhibitory effect on proliferation of cancerous cells and their anti-
proliferative effect is fully blocked by histamine -4 receptor antagonists like JNJ7777120 [5,34].
Administration of histamine-4 receptor antagonists have both anti-inflammatory and anti-pruritic
effects and increases level of astrocytes, release of inflammatory cytokines, increase microglia
migration and activation in the CNS which are critical for the demyelination and for clinical
signs of encephalitis [28]. Histamine -4 receptor antagonists have the ability to inhibit the effect
of histamine and 4-methyl histamine induced calcium mobilization and Th2 cytokine IL-13
release by inhibiting the activation of ERK1/2, Akt and NF-kB when stimulated with histamine

26
in human mastocytomacells-1 [6, 35]. Even though H4R antagonists exhibited inhibited allergic
inflammation, with decreases in Th2-mediated IL- 4, IL-5, IL-13 and IL-17 in several
experimental models Conversely, stimulation of H4R agonist down regulates the production of
Th1-mediated cytokines including IFN-a, TNF-a, IFN-c, IL-12 and IL-27 on certain subtypes of
dendrite cells involved in inflammatory skin diseases. For instance, the activation of H4R
agonist like 4-methyl histamine might play a role in the modulation of pro-inflammatory immune
responses of allergic skin diseases, especially in Th1-mediated pathologies such as psoriasis [30,
36].

27
4. Future perspectives

Future research in the treatment of inflammatory and autoimmune disorders should search for
new therapeutic approaches that accomplish complete resolution of symptoms while approaching
simultaneously all tissues where an inflammatory response is evident (mainly respiratory tract
and skin), as well as enhancing patient compliance and comfort. In an ideal world, these should
be etiological approaches to new pharmacological targets with appropriate safety profiles,
allowing the use of systemic routes of administration that provide an integrated approach to
inflammatory and autoimmune disorders as a whole. Still now there is strong preclinical
evidence on H4R as a promising target for the treatment of those disorders. Unlike other
histamine receptors, its preferential distribution pattern in immune cells such as mast cells,
eosinophils, dendritic cells and T cells, suggest an important role of H4R in the immune
response. In vitro studies have confirmed an important role of H4R in cellular responses,
including chemo taxis, Th2 polarization, and cytokine and chemokines production. In vivo
studies in different species using several histamine H4 agonists and antagonists have confirmed
the potential of H4R modulation in inflammatory and autoimmune disorders, although species
differences regarding compound’s behavior i.e. agonist/antagonism, have raised some conflicting
results. Currently several H4R antagonists have already initiated clinical trials, both in healthy
volunteers and in patients with inflammatory and autoimmune disorders. Results from the
ongoing clinical studies are excitedly appointed to establish the potential role of H4R antagonists
in the treatment of inflammatory and autoimmune disorders.

28
5. Conclusion and recommendations

Histamine is an important chemical messenger that exhibits significant physiological effects

mediated through its receptors. A thorough knowledge of drugs is very much useful to treat the
clinical conditions arising due to imbalance of histamine in the body. H1 receptors act on the
gene transcription of inflammatory mediators. H2 antagonists are practically non-sedative
because it is not permeable through BBB. When a person has a problem in metabolizing and
breaking down histamine, its level build up and lead to worrying symptoms like rashes, itching,
swelling, redness etc. All who have histamine intolerance in any kind need to take steps and
manage it by taking specific diets and supplements such as protein- dairy products, egg whites,
peanuts; vegetables –tomatoes (red one has higher histamine than white or yellow one), spinach,
pumpkin etc. Despite of the fact that the majority of the H1-antihistamines have similar efficacy
in the allergic diseases, after extensively research, it can conclude that the second generations of
antihistamines constitute the medication of choice for the treatment of allergic rhinitis, allergic
conjunctivitis and chronic urticaria. The low histamine diet may help people who develop
symptoms, such as sneezing, itching or hives, in response to foods that contain histamine.
Histamine is a chemical that occurs naturally in the body and some foods. The low histamine diet
can help a person find out which foods cause their symptoms. By avoiding those foods, they may
see an improvement.

29
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