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Gene transcription is important in regulation of cell function, it is medaited by a number of reactions done on amino acids of lysine tails in histone protein which are responsible for DNA packing and inhibition of gene transcription. One of these reactions are involved in the addition or removal of acetyl groups known as acetylation and deacetylation which alter the DNA structure thus altering gene transcription. Histone deaceylase (HDAC) is the enzyme responsible for deacetylation which is the removal of acetyl group to maintain histone binding to DNA thus reducing gene transcription. Targeting HDAC has been attractive as a therapeutic method in diseases, in particular a variety of HDAC inhibitors have been developed for cancer. Cancer is a major disease involving abnormal cell growth regulated by gene transcription. However it is also known that various inflammatory diseases are also related to gene transcription levels and there has been growing interest in targeting HDAC as a therapy method. This review will focus on the use of HDAC target therapy in a variety of inflammatory diseases such as sepsis, rheumatoid arthritis, multiple sclerosis, systemic lupus erythematosus, traumatic brain injury, haemorrhagic shock and Huntingdon disease. This is in order to assess if HDAC targeted therapy is a beneficial and ideal in the treatment of inflammatory diseases.
Cells are the basic building blocks of an organism; they carry out various functions such as division, migration or release of mediators. In order to carry these functions they require messengers or signals in the form of gene transcription. Gene transcription in itself is regulated by modification of the DNA nucelotide bases of genes in forms of reactions such as methylation, acetylation and deacetylation. This modification results in the alteration of gene function such as increase or decrease of gene activity.
The HDAC enzyme is responsible for deacetylation which is usually involved in reduction of gene transcriptions. As they are involved in many cell processes, deregulation of these enzymes can be related to a variety of diseases. One of these diseases is cancer in which is caused by a lack of acetylation (thus reduction in gene transcription). HDAC inhibitors are being developed as cancer therapies.
Another disease of interest is those around inflammation, since pathology between common inflammation and cancer are similar and are dependent on gene transcription. There has been interest in designing therapies targeted towards HDAC in cancer treatment. Could the same be said for HDAC targeted therapy towards various inflammatory diseases? This review will cover HDAC involvement in various inflammatory diseases to assess if HDAC is a potential target for therapy of these diseases.
3. DNA and nucleosome arrangement
The total amount of DNA within a human is long. In average the DNA length within a chromosome is around 19,000µm to 73,000µm, thus the total length of DNA within 46 human chromosome is at 2 metres long (William et al 2007). This length of DNA needs to be restricted within the nucleus of a cell with a diameter of 5 to 10µm. DNA is packed into the nucleus is carried out through the help of histones, proteins in which DNA is wounded up around against to form a nucelosome. A nucelosome consists of 146 base pairs of DNA wrapped around the core histone octamer (Marks et al., 2001). Nucleosome forms the basic unit of a chromosome which can be seen in the nucleus when condensed. The core histone octamer consists of 2 pairs of histones H2A, H2B, H3 and H4 (Timmermann et al., 2001). Figure 1 shows the DNA arrangement into a chromosome (taken from William et al 2007).
Figure 1: DNA organisation within the cell (William et al 2007).
The 4 main histones contain an amino acid terminal tail consisting of the C and N terminals; these are lysine rich residues that are charged. Under normal physiological conditions the residues are kept positively charged. The N-terminal tail of the amino acid side passes through and around the DNA double helix structure. Negative charges within the phosphate group on DNA bases interacts with positive charges on histone amino acid terminals resulting in the tight formation of the nucleosomes (Timmermann et al., 2001). It is at the C and N terminals known as the lysine tails where its structure can be altered through reactions of acetylation or deacetylation. This is important for modulation and regulation of gene transcriptions. Figure 2a shows the structure of the histone core, followed by the amino acid sequence of the lysine tails shown in 2b (taken from Marks et al., 2001).
4. Acetylation and deacetylation
Acetylation reaction involves the addition of acetyl groups to the lysine tails of the histone at the C or N terminal. This results in neutralization of the charges between DNA bases and histone which weaken the association between each other, this can be demonstrated in testing acetylation in histone H4 binding to DNA in which addition of acetyl groups resulted in weak interaction between the two (Hong et al., 1993). As a result of acetylation the DNA is exposed to transcriptional factors to bring a variety of cellular responses and functions though gene transcription. Acetylation reactions are mediated and initiated by histone acetylate enzymes (HAC), the end result is increased gene transcription.
The opposite of acetylation is deacetylation which is the removal of acetyl groups at the histone lysine tails. This as a result leads to the restoration of the positive charges on the lysine histone tails to reform the strong interactions between the DNA bases and the histone protein. Overall the end result is the reconstruction of the nucelosome that prevents gene transcription. Deacetylation reactions are mediated and initiated by histone deacetylates enzymes (HDACs).
Figure 2a: structure and composition nucelosome organisation consisting of histone protiens. Figure 2b :arrangement of amino acid in lysine tails of the histone proteins (Marks et al., 2001).
5.1 HDAC- Types
The HDAC family is currently organised into 5 classes and spilt into 2 types depending on structure and mechanism. In general the first type of class is the zinc dependent group consisting of classes I, IIa, Ilb and IV. The second type of class is the non zinc independent class III.
Zinc dependent HDAC have a zinc atom responsible for its catalytic activity. Class 1 consists of HDAC members1, 2, 3 and 8, which are located within the nucleus. They consists of a nuclear localization signal, in addition HDAC 3 has a nuclear export signal. Class 2 have HDAC members 4, 5, 7 and 9. These proteins consist of extended amino acid domains of 600 amino acids along with a zinc catalytic domain (Kazantsev et al., 2008). Class 2b are structurally different to class 2a HDACs with HDAC members 6 and 10. HDAC 6 has 2 independent catalytic domains in addition to a zinc finger ubiquitin binding domain, while HDAC 10 lacks one of the catalytic domains. Finally class 4 contains HDAC member 11, this is located within the nucleus, however not much is known about this member. Expression is high within the brain, heart, muscle, kidney and testis. Investigations of this protein have been identified to have similar structure to HDAC member 6 in co-precipitation reactions (Gao et al., 2002).
As for non zinc dependent HDAC, This group only consists of class 3, these set of protein are structurally and functionally distinct from the other HDACs. These members are named the silent information regulator 2 (Sir2) based on its discovery in budding yeast, there are 7 members. They are dependent on NAD+ for their catalytic reactions such as deacetylation activity and ADP transferase (Kazantsev et al., 2008). Table 1 summarizes the HDAC family with drugs targets.
Table 1. HDAC classification
Drugs that affect HDAC type
SAHA, TSA, Butyrate, Theophylline, Glucorticoid
SAHA, TSA, Butyrate
Non Zn2+ dependant
5.2 HDAC mechanism of action
This mechanism of lysine deacetylation on DNA bound histone by HDAC is not understood fully, thus this is a proposed mechanisms of HDAC by Finnin.
For zinc dependent group HDAC, this is suggested to start at intermediate chelating of the necessary water molecule and the carbonyl group of the acetyl lysine to the zinc atom (Bertrand, 2010). It is then assumed that histidine 132 is protonated, this leads the water to be deprotonated by histidine 131 which results in the protonation on histidine 131 (Bertrand 2010). This forms a hydroxide ion, which is able to attack the carbonyl of the acetyl group (Bertrand 2010).
However there have been contradictions in which other proposals of HDAC mechanism that have been suggested. One example is histidine 131 requirement of tyrosine 297 aid to form a hydrogen bond to produce the hydroxide ion (Vanommeslaeghe et al., 2005). In addition, there have been objection to the formation of the hydroxide ion due to mismatch of negative charges formed overall, another proposal suggests a nucleophilic attack of a water molecule, which is facilitated by histone 132 (Corminboeuf et al., 2006).
As for the mechanism involving the non zinc dependent group HDAC sirtuin, is similar but different to zinc dependent HDACs. Sirtuin binds on to the acetyl group of the lysine tail, followed by cleavage of Nicotinamide adenine dinucleotide (NAD+). This produces nicotinaminde and an alkylamide intermediate which attacks the carbonyl oxygen of an acetyl group. The end result is production of ribose and the deacetylated production (Denu, 2005).
5.3 HDAC functions
HDACs don't have the ability to bind on to DNA; instead they recruit a large number of genes through direct association with transcriptional activators and repressors (Haberland et al., 2009). The selectivity and function of HDACs depends on a number of factors including cell identity, protein members available in the cell and the signalling environment of the cell (Haberland et al., 2009).
HDACs have been known to be associated with cell cycle regulators such as Mad/Max which are transcription factors of a gene named c-myc originating from a cell line of a retrovirus c-mcy-ocogene (Dang, 1999). Mad/Max heterodimers are responsible for the inhibition and repression of E-box consisting of genes for growth stimulatory during cell differentiation, which are affected, in the pathology of basic cancer (Dang, 1999).
Apart from this not much is known about HDAC exact influence outside gene regulations such as protein binding.
5.4 HDAC inhibitors and simulators
There has been a range of HDAC inhibitors mainly designed towards initially for cancer, but now have been looked as potential therapies for various inflammatory diseases. There are 3 types of HDAC inhibitors short chain fatty acids, hydroxamic acid and non classified types (Marks et al., 2001).
Hydroxamic acid types such as Suberoylanilide hydroxamic acid (SAHA) inhibits class II and I of HDAC through binding of the zinc pockets of HDAC. A non-classified HDAC inhibitor is Trichostatin A (TSA) which inhibits class I and II of HDAC like SAHA. Fatty acid type Butyrate inhibits all class I and II HDAC members apart from HDAC 6 and 10 (Blanchard et al., 2005).
There are HDAC simulators such as Glucorticoids which enchance activity of HDAC 2 (Ito et al., 2000) and Theophylline which enhance the activity of HDAC 1 (Ito et al., 2002).
6. HDAC and cancer
Although cancer is not an inflammatory disease, is it a critical disease involving abnormal cell division growth affecting many people around the world. One of the potential key mediators of cancer is those surrounding HDAC.
Cancer is a major disease affecting many people throughout the world. Currently in the United States a total of 1,529,560 new cancer cases and 569,490 deaths from cancer are predicted to happen (Jemal et al., 2010). Cancer worldwide in 2002 has caused 10.9 million affected people, 6.7 million deaths, and 24.6 million cancer survivors (Parkin et al., 2005). The most common cancer types is lung cancer (1.18 million deaths), stomach cancer (700,000 deaths), and liver cancer (598,000 deaths) with breast cancer being the most preventable and treatable (4.4 million) (Parkin et al., 2005).
It is known that a range of HDAC is involved in the pathology of serial cancer types. Such as HDAC1 and HDAC2 over expression in prostate, gastric and colorectal cancer, as well as HDAC3 over expression in lung cancer (Bertrand 2010). However there has been low expression of class HDAC5 in colon cancer and acute myeloid leukaemia (Bertrand, 2010).
7.1 HDAC targeted therapy in models of inflammatory diseases
Inflammatory diseases are mainly related to a deregulation of genes resulting in major symptoms such as pain, redness, swelling and elevated body temperature.
Recently, HDAC have been attractive drug targets in inhibition as well as in stimulation mechanisms for the treatment of inflammation, since the basis of inflammatory pathology in terms of response is mediated through gene transcription.
Sepsis is a systematic inflammatory disease resulting in multiple organ failure resulting death in many hospital patients. A study was conducted in United States between 1999 and 2005 to observe the affects of sepsis clinical patients; the results show that 1,017,616 patients out of 16,948,482 deaths were related to sepsis (Melamed et al., 2009).
Sepsis is a complex disease in which its pathology consists of many components and mediators such as NF-Ä¸B, cytokines TNF-Î± and leukocytes neutrophils and macrophages. It seems that sepsis is related through the deregulation of genes, in which HDAC inhibitor have been effective in treatment of sepsis models in vitro studies.
SAHA administrated to mice injected with endotoxin LPS, resulted in the inhibition of inflammatory cytokine levels of TNF-Î±, IL-1Î², IL-6 and interferon gamma (IFN-Î³) (Leoni et al., 2002). Similarity, administration of TSA demonstrated reduction in hepatic injury biomarkers AST and ALT, lower myeloperoxidase (MPO) activity and suppressed expression of intercellular adhesion molecule-1 (ICAM-1) in liver tissue, when administrated in CLP induced mice (Zhang et al., 2009).
However in comparison, simulation of HDAC is said to be effective in improving septic condition in particular in muscle wasting which is a common feature in sepsis. It has been identified that septic rats have reduced muscle breakdown through increased levels of p300 and HDAC activity suggesting HDAC enhancers can improve muscle condition induced by sepsis (Alamdari et al., 2010). Other contraindications is the administration of the HDAC inhibitors SAHA and TSA increased LPS-induced expression of interleukin 6 (IL-6) and inducible nitric oxide synthase mRNAs. As well as the secretion of cytokines IL-6, TNF-Î±, macrophage inflammatory protein 2 (MIP-2), and nitric oxide (NO) in murine N9 and rat primary microglial cells. It was suspected that nucleur factor kappa beta (NF-KB) amplification by LPS induction is involved in one half of cytokine and NO simulation that is independent of TSA administration indicating that HDAC has a role in affecting inflammation response through gene transcription (Suuronen et al., 2003).
7.3 Rheumatoid arthritis
Rheumatoid arthritis is a common chronic inflammatory disease mostly affecting synovial joints. Synovial joints are the most common type within the human body, responsible for connection and movement of articulating bones within hands, fingers and legs. In a clinical study involving 7050 individuals in Norfolk, UK, 66 cases of that population sample obtained RA symptoms. 1.16% of the cases were woman while 0.44% was men (Symmons et al., 2002).
Patents that have rheumatoid arthritis commonly have pain and stiffness in multiple joints. This pain and stiffness is restricted to one joint only but spreads and develops weeks or months later. Additional symptoms include anorexia, weakness, or fatigue (Rindfleisch et al., 2005). The disease seems to be induced by a number of factors. In particular female sex, a positive family history, older age, silicate exposure, and smoking are high risk factors which might contribute to the disease (Rindfleisch et al., 2005).
A common synovial joint consists of the articulating bones connected via a synovial capsule (joint capsule) containing synovial fluid. The end of the articulating bones are coated with articular cartilage, both the articular cartilage and synovial fluid prevent unnecessary friction of the two articulating bones during movement.
In joint rheumatoid arthritis, various components and mediators of inflammation such as cytokines TNF-Î±, IL-1, IL-6 and leukoctyes of macrophages are primed and attack the synovial joint. As a result bone is eroded as well as the articular cartilage is broken down and lost. This concludes in frequent articulate bone friction causing pain during movement at the affected joint.
However the actual mechanism of initiation of inflammation at the synovial joint is still unclear. In general there is activation in proliferation of synovial macrophages and fibroblasts after an unknown triggering incident. Later on in the course of the disease, the inflamed synovial tissue begins to grow irregularly, forming invasive pannus tissue which invades and destroys cartilage and bone. This destruction is amplified by multiple cytokines, interleukins, proteinases, and growth factors (Rindfleisch et al., 2005).
These levels of inflammatory mediators and components are induced and control via expression of gene transcription through various pathways such as NF-ÎºB, mitogen-activated protein kinase (MAPK), phospha-tidylinositol 3 kinases (PI3Ks), Janus tyrosine kinase (JAK) and activators of transcription (STAT) pathway.
There has been evidence of increased HDAC levels in synovial samples from rheumatoid arthritis joint patients suggesting a role of HDAC in transcriptional regulation of pro-inflammatory genes (Huber et al., 2007). SAHA administrated to rat collagen induced arthritis (CIA) models reduced symptoms of paw swelling, bone erosion and onset of arthritis. However the drug does no inhibit the onset of the disease (Lin et al., 2007a).
7.4 Multiple sclerosis
Multiple sclerosis is an autoimmune disease of the nervous system. Where the fatty myelin sheaths around the axons of the brain and spinal cord are damaged. This results in demyelination of axons and disrupts communications between nerve cells in the brain and spinal cord to communicate with each other. Without communication basic physical processes such as movement are disputed.
In epidemiological studies of multiple sclerosis cases in Europe. The estimate of MS State is 83 per 100,000 populations with higher rates in northern countries. In which females are more susceptible to the disease then males (Pugliatti et al., 2006).
Clinical symptoms of multiple sclerosis are assessed by expanded disability status scale (EDSS) (Kurtzke, 1983). Here 8 sections of assessment called Functional Systems (FS) these are pyramidal, cerebellar, brain stem, sensory, bowel/ bladder, visual, cerebral, and other. Each of these sections is rated from 1 to 10. This is calculated into a total score of disability status scale (DSS) which is also scored from 1 to 10 (Kurtzke, 1983). Another diagnosis is the detection of the number silent lesions in the brain through magnetic resonance imaging (MRI). In addition of number of "clinical attacks" observed in patients.
The inflammatory pathway of the disease involves initial silent lesions of the blood brain barrier (BBB) of the nervous system, which are caused by initiation of CD4+ helper type cells (TH1 cells) (McFarland et al., 2007). In addition there is also a high abundance of activated macrophages or microgia involved in the inflammatory process (Lassmann et al., 2007). The silent lesion develops over a timespan of a month and then is repaired through re-myelination of some axons but this leaves area of damage (Lassmann et al., 2007).
Levels of cytosolic HDAC 1 were detected in damaged axons of mice with cuprizone-induced demyelinationone a experimental model of multiple sclerosis and damaged axons in brains of humans with multiple sclerosis, suggesting HDAC involvement in the pathology of axon degeneration (Kim et al., 2010). Histone deacetylase (HDAC) inhibitor drug TSA has demonstrated reduction in spinal cord inflammation, demyelination, neuronal and axonal loss when administrated another experimental model of multiple sclerosis known as autoimmune encephalomyelitis (EAE) (Camelo et al., 2005).
7.5 Systemic lupus erythematosus (SLE)
Systemic lupus erythematosus (SLE) is another autoimmune disease that is characterized through clinical implications of multisystem organ function and immunological symptoms of antinuclear antibody production (Lipsky, 2001).
In an American study done between January 1, 1980 and December 31, 1992. 430 medical patient's studies were analysed. Within the 430 patients 48 of the patients have the SLE condition, 42 women compared to 6 men (Uramoto et al., 1999).
Women are mainly affected by the disease, along with factors of genetic susceptibility, gender, stress and environmental (Lipsky, 2001). There is also genetic susceptibility of SLE is due to a variety of gene variation, one is gene variation on chromosome 6 of HLA class 1 and 2 (Martens et al., 2009). Other genes suspected to be involved is STAT4 and BLK but this is related more to the population in Hong kong (Yang et al., 2009).
SLE is characterized by a high level of cytokines produced by t-cells/b-cells activation, deregulated antibody production and renal inflammation. There is suggestive production of cytokines production of IL-6 and 10, in addition to high production of IgG antibody production all produced from b cells (Lipsky, 2001).The mechanism and pathology of the disease is still not understood, one suggestion is the alteration of gene transcription, which regulates inflammatory response.
Administration of HDAC specific inhibitors TSA and SAHA in an SLE model of MRL-lpr/lpr mice, demonstrated reduction of IL-6, 10, 12 and IFN-Y production from splenocytes (Mishra et al., 2003).
7.6 Traumatic brain injury (TBI)
Traumatic brain injury (TBI) is not an inflammatory disease but does activate inflammatory pathways to develop into an inflammatory disease. It is caused by extensive damage to the brain that causes consequences such as axonal injury, cell death, contusions, and inflammation.
In America it is estimated that TBI incidence is 180 and 250 per 100,000 populations per year. In particular the young males are affected. Mortality and severity rate varies in patients, but rates are high in those with severe injury and in the elderly (Bruns et al., 2003).
Early symptoms of traumatic brain injury include headache, dizziness, nausea, vomiting, slurred speech, imbalance, and incoordination. This also concludes signs of confusion that includes vacant stare, disorientation, delayed verbal or motor responses, and poor concentration or attention (Kushner, 1998).
The inflammation response is rapid and robust, characterized by the release of cytokines and activation of microgila. Cytokines that could be released through the microgila is IL-la, IL-1P, IL-6, TGF-Î±, TGF-3, IFN-Î±, IFN-3, IFN-y, TNF-Î±, G-CSF and GM-CSF. Macrophages could also be activated through cytokine activation and production (Ghirnikar et al., 1998).
In a rat model of TBI, there was decreased levels of hippocampal CA3 histone H3 acetylation 72 hours after injury, suggesting HAT is has a role in the pathophysiology of TBI (Gao et al., 2006). A novel histone deacetylase (HDAC) inhibitor DMA-PB was administrated in rat models of TBI, the results demonstrated an increase in levels of HAT 3 as well as reducing phagocytic microglia within the Hippocampal region (Zhang et al., 2008).
7.7 Haemorrhagic shock
Haemorrhagic shock is similar to TBI but not restricted to the brain. This is a condition where there is a substantial loss of blood mainly following extensive damage.
In America it is estimated that TBI incidence is 180 and 250 per 100,000 populations per year. In particular the young males in particular are affected. Mortality varies by severity varies but is high in those with severe injury and in the elderly (Heckbert et al., 1998).
Loss of blood flow following blood loss results in a reduction in blood circulation throughout the body and organs causing hypoperfusion and dysfunction within resulting in shock. Blood is a critical component of the body responsible for the delivery of oxygen and nutrients to various cells which affects the central nervous, cardiac, and renal systems.
Hypoperfusion of organs forces cells to go under anaerobic metabolism resulting in acidosis. It is the production of tissue hypoxia, acidosis and various mediators cause activation of the inflammatory response which is systemic. This can result in injury of cells or tissues from released oxygen radicals, activation of the humoral and cellular inflammatory systems that can inflict vascular and cellular injury. During shock the body is prone to infection by various micro-organisms which can result in systemic inflammatory response syndrome (SIRS) and multiple organ failure.
It is not certain on what inflammatory components are involved in the pathology of hemorrhagic shock. There is suggestion of an increase of TNF-Î± and IL-6 in septic and trauma patients with hemorrhagic shock (Martin et al., 1997). Other inflammatory mediators involved are nitric oxide and neutrophil activation (Hierholzer et al., 1998).
Rats that underwent hemorrhagic shock were found to have partial levels histone deacetylation, fluid resuscitation using ketone ringer's solution demonstrated hyperacetylation of histones H2B, H3, and H4. Suggesting balance of HDAC/HAT activity is related to haemorrhage and resuscitation (Lin et al., 2006). This theory was supported in another investigation in which administration of 3 other HDAC inhibitors, VPA, TSA, and SAHA in fluid resuscitation demonstrated results of an increase in acetylation activity similar to rats undergoing fluid resuscitation (Lin et al., 2007b). Finally administration of SAHA to hemorrhagic shock induced rats, demonstrated TNF-Î± reduction suggesting HDAC is beneficial in reducing inflammatory responses in hemorrhagic shock (Sailhamer et al., 2008).
7.8 Huntingdon disease (HD)
Huntingdon disease is a progressive neurodegenerative genetic disorder, which causes defects in muscle coordination and development of memory dysfunction. It is caused mainly through a genetic inheritance of a mutant of huntingtin, which encodes for multiple CAG repeat on the first exon causing dysfunction to the protein.
A retrospective trail in England was taken place between 1987 and 1990 in genetic centres. Overall 248 subjects were in risk of Huntingdon disease and were predicted. Out of the 248 patents 151 (61%) of the applicants were not in risk of the disease, while 97 (39%) were in risk of the disease. 158 (64%) of the subjects were female and 90 (36%) male (Tyler et al., 1992). Another worldwide study of huntingdon disease concluded that the disease is highly common in European populations, both Northern and Southern, through a relatively high prevalence rate (4-8 per 100,000 population) (Harper, 1992).
Symptoms of Huntingdon is very hard to detect in the early stages of the disease, it is not until the later stages of the disease that clinical signs can be detected. This is usually in middle-age after affected individuals have had children. Late symptoms include cognitive dysfunction such as long-term memory and impairment of executive functions, such as organising, planning, checking, or adapting alternatives, and delay the acquisition of new motor skills. These conditions worsen as time progresses in particular speech. Other symptoms are also behavioural like depression and suicidal thoughts (Walker, 2007).
Within the pathology of Huntingdon disease, there have been suggestions in activation of the immune system during development of Huntingdon disease. Plasma samples collected from HD carries ranging from premature and mature states. The plasma was found to have high levels of cytokines IL-6, IL-8, IL-4, IL-10, TNF-Î±, IL-5, IFN-Î³ and GM-CSF in premature and mature states of HD patients than normal patients .
HDAC have been beneficial in various models of Huntingdon disease, sodium butyrate administrated to models of R6/2 transgenic mice a genetic model of HD increases body weight, motor performance and survival rate as well as increasing neuroprotection rate (Ferrante et al., 2003). In addition HDAC inhibitor SAHA administrated orally to HD model mice improved symptoms of motor defects. It also identified that SAHA can across the blood brain barrier to increase histone aceylation in particular histone H2B and H4 within the brain (Hockly et al., 2003).
Stroke is a condition of brain dysfunction which is caused by interruption of brain blood flow through loss of blood flow known as ischemia, blockage or blood leakage known as haemorrhage. In the past the disease was termed as cerebrovascular accident (CVA).
A analysis of worldwide deaths in 1997 showed that stroke is the second highest cause of global deaths behind ischaemic heart disease with 4·38 million deaths in developed countries and almost 3 million in developing countries (Murray et al., 1997). In western countries this proportion of death is 10-12%, this percentage consist of people that are below the age limit of 65 years old (Bonita, 1992).
Normally stroke is clinically diagnosed through the use a number of scanning equipment such as computed tomography (CT) and magnetic resonance imaging (MRI). In the past CT have been used for diagnosis but now the introduction of MRI it has been to be used instead due to the ability of MRI to accurately detect pathological symptoms of stroke patients in particular in suspected acute stroke (Chalela et al., 2007).
For non diagnosed classification in various countries several systems have been proposed and recommended for quick diagnosis of suspected stroke patients, in the UK this is FAST (face, arm, speech, and time) (Harbison et al., 1999). In America this is Los Angeles Prehospital Stroke Screen (LAPSS) (Kidwell et al., 1998) and Cincinnati Prehospital Stroke Scale (CPSS) which consists of detecting 3 main physical symptoms of the patients arm weakness, speech, and facial droop (Kothari et al., 1999).
The pathophysiology of stroke is complex, and involves excitotoxicity mechanisms, inflammatory pathways, oxidative damage, ionic imbalances, apoptosis, angiogenesis and neuroprotection (Deb et al., 2010). In the case of ischemic brain stroke there are 3 subtypes of manifestation in the form of thrombotic stroke (large vessel and small vessel types); embolic stroke (with/without known cardiac and/or arterial factor); systemic hypoperfusion (Watershed or Border Zone stroke) and venous thrombosis (Deb et al., 2010).
Ischemia can be caused by a number of factors in particular atherosclerosis which is formation of a plaque in the blood vessel following injury and inflammatory response restricting blood flow of the vessel. The various cytokines and inflammatory components detected in the onset of stroke involved adhesion molecules of ICAM, VCAM-1 and selectin, as for cytokines suspected to be involved are interleukin-1 beta (IL-1Î²), interleukin-1 receptor antagonist (IL-1ra), IL-6, interleukin-8 (IL-8), interleukin-10 (IL-10) and tumour necrosis factor alpha (TNF-Î±) (Deb et al., 2010). This results in an end point of reduced blood flow restriction of blood and glucose supply to brain cells, causing cell death thus reduction in brain function.
From investigations there are suggestions that heat shock protein 70 (HSP-70) is neuroprotective against ischemic induced strokes (Kinouchi et al., 1993). The suspected mechanism of HSP-70 neuroprotection is assisting correct protein folding in regulation of cells thus preventing risk of cell death (Giffard et al., 2004). Thus biased on this theory, administration of HDAC inhibitor VPA reduced brain damage and increase functional outcome of rats induced via transient focal cerebral ischemia. The suggestion of this attenuation is the over expression of HSP-7 activity (Ren et al., 2004). Similar findings were demonstrated in the administration of SAHA increased expression of HSP-7 to attenuate ischemia brain area of mice after ischemia damage (Faraco et al., 2006).
Finally an intensive investigation of administration of various HDAC inhibitors VPA, butyrate, or TSA had benefits of treatment in a rat permanent ischemic model of stroke. Administration of the inhibitors demonstrated a reduction in brain infract size, in addition VPA or butyrate suppressed microglial activation and reduced the number of microglia, and inhibited other inflammatory markers in the ischemic brain. Once again this is expected to be related to the increased levels of HSP-7 activity (Kim et al., 2007).
8.1 HDAC in clinical trails
Since the development and successful experiments involving HDAC and various disease models, it is natural that HDAC will be tested in the actual disease state in humans carried out by clinical trials. However only a few inflammatory diseases have HDAC tested in the relative clinical trials. The majority of HDAC clinical trials have been more towards the cancer field.
8.2 HDAC in clinical trials- cancer
For cancer clinical trials Vorinostat also known SAHA have been tested in various clinical trials, one includes clinical trial phase 1 T-cell lymphoma in which a few side effects of diarrhoea, fatigue, nausea and anorexia (Olsen et al., 2007). Romidepsin is another HDAC which has a sulfhydryl group which functions as a zinc chelator, a phase 1 clinical trial tested Romidepsin in patients with refractory neoplasms, a range of side effects were found in the form of fatigue, nausea, vomiting, cardiac arrhythmia and thrombocytopenia (Sandor et al., 2002).
As for inflammation diseases, the only two inflammatory diseases in which clinical trials of HDAC targeted therapy have been successful in patients are ulcerative colitis and COPD.
8.2 HDAC in clinical trials of inflammatory disease- Ulcerative colitis
Ulcerative colitis is a form of inflammatory bowel disease, the large intestine or colon is the area affected. It is characterized by ulcers and open sores.
To assess the rate of the disease, a study of ulcerative colitis patients was conducted throughout 20 European centres was conducted between 1 October 1991 to 30 September 1993. Out of 2201 patients aged 15 years or more, 1379 were diagnosed to have ulcerative colitis. Iceland was the country to have the highest rate of ulcerative colitis (Shivananda et al., 1996). There also seems to be a genetic relationship in various 12 regions in chromosome 16, 12, 6, 14, 5, 19, 1, 16, and 3 in ulcerative colitis patients (Baumgart et al., 2007).
The inflammatory pathway is not well known, however it is suggested that a over reactive disease which causes damage to the tissue. There is a suggestion that inflammatory mediator NF-Ä¸B is mainly involved. Also there are additional inflammatory mediators involved such as t cells such TH-17 activation, TLR-4 increased expression, nitric oxide, oxygen radicals, prostaglandins, leukotrienes, histamine, proteases, and matrix metalloproteinases (Baumgart et al., 2007).
HDAC inhibitor brutyate has been successful in clinical trials of ulcerative colitis patients. One trail demonstrates butyrate given tropically to patients improve symptoms of the disease via increased bowel movements (Vernia et al., 2003). While in another clinical trail brutyate given for 4 to 6 weeks in patients had reduced NF-KB levels within macrophages (Luhrs et al., 2002).
8.3 HDAC in clinical trials of inflammatory disease - COPD
COPD is an inflammatory disease of the lung, where lung parenchyma and emphysema are destructed resulting in irreversible narrowing of small airways.
An investigation of UK 50,714 COPD patients were analysed in conditions of COPD from 1990 to 1997. The result demonstrated 23,277 (45.9%) of the COPD patients were female, over the years the rate of women diagnosed with COPD increased from 0.80% to 1.36%. Indicating that the death rate is higher in women then male, the mean age range of these patients was 76.5 years (Soriano et al., 2000).
Lung function is measured by the ability of the lung being able to expel air in a second this measured as FEV1. According to GOLD (global initiative on obstructive lung disease) FEV1 can be used to diagonalize to assess various states of COPD (mild, moderate and severe) (Pauwels et al., 2001). For COPD patients the FEV1 declines as age increases, figure 4 shows decline of FEV1 in COPD state patients as well as improvement of FEV1 if COPD stopped smoking (Hogg, 2004).
The pathology of COPD involves 2 inflammation pathways of the airways known as chronic bronchitis and emphysema. The chronic bronchitis happens in the epithelium of the central airways, where there is an increase of mucus production with disruption of the epithelial cell wall by innate and immune cells. Emphysema involves the destruction of the alveoli wall, and is related to the smoking a common factor in COPD patients. It is not certain what the exact pathology underlying this mechanism.
Gene transcription is responsible for mediating these inflammatory conditions, as well as smoking being the most common factor for inducing the inflammatory process within the lung. There is evidence of reduced HDAC levels in patients with increasing progression of COPD, suggesting that HDAC is a key gene transcription repressor in inhibiting proinflammatory cytokines in alveolar macrophages which are involved in inflammation of COPD (Ito et al., 2005).
Glucorticoids such as dexamethasone are known to active HDAC in particular HDAC 2 to reduce proinflammatory genes like IL-1Î² stimulated granulocyte-macrophage colony-stimulating factor in adenocarcinomic human alveolar basal epithelial cells (A549 cells). So in theory glucorticoid should be an effective treatment in COPD patents (Ito et al., 2000). However in clinical studies administrating glucorticoids to COPD patents demonstrates no improvement in symptoms or conditions as well as no anti-inflammatory effects (Culpitt et al., 1999).
Theophylline is a HDAC activator. When administrated in alveolar macrophages from COPD patients in a clinical trial, Theophylline increased HDAC expression in conjunction with glucorticoid dexamethasone suggesting Theophylline can be used to improve COPD condition along with glucorticoid (Cosio et al., 2004). In addition Theophylline has been tested in a small clinical study of 30 COPD patients administrated in conjunction to corticosteroid fluticasone propionate. The results demonstrated reduction in eosinophils and increases in forced midexpiratory flow rate and FEV improving COPD symptoms of these patients (Ford et al., 2010).
Figure 4 classification of FEV1 state in COPD patents (Hogg 2004).
9.1 HDAC potential therapy in inflammatory disease
HDAC discovery in the pathology of inflammatory diseases is still active and on-going. This indicates that HDAC therapy can still have potential in therapeutic treatment of other inflammatory diseases not yet tested in experimental models.
9.2 Hepatitis C (HVC)
Hepatitis C (HVC) is a strain of virus that infects liver to cause a variety of liver or other related diseases such as cirrhosis, digestive tract haemorrhage, liver failure, and liver cancer (Poynard et al., 2003). It is a positive strand RNA virus which is related to flavi and pestiviruses family (WHO 1999). The discovery of HVC was in 1989 through the isolation of its cDNA from a patients with NANBH (non A, non B hepatitis) (Choo et al., 1989).
From world health organization (WHO) estimates it is predicted that approximately 3% of the world population or about 170 million people, may be infected with hepatitis C virus (1999). Diagnosis of the disease is mainly through serological assays for antibodies and molecular tests for viral particles. For serological tests, antibodies of the virus are detected through enzyme immunoassays; this can take 4-10 weeks after infection. While the detection of viral particle is through assay identification of activity in Hepatitis C viral RNA (Poynard et al., 2003).
The virus induces an immune response to cause destruction of hepatic cells. This is suspected through the activation of CD4+ cells, which was indicated and activated in high levels of liver biopsies and peripheral blood mononuclear cells (PBMC)of chronic hepatitis C-infected patients in response to protein nonstructural 4 (NS4), a recombinant form of HVC (Minutello et al., 1993). Another important inflammatory mediator is production of reactive oxygen species from oxidative stress to influence another important non inflammatory mediator of the disease which is iron. Iron overproduction within the liver is toxic; this seems to be a main factor present in Hepatitis C infection. It seems that iron availability is regulated by a key molecule hepcidin in which its activity is suppressed by reactive oxygen species (Nishina et al., 2008).
HDAC inhibitors TSA and PDTC increased hepcidin levels in HVC infected cells (Miura et al., 2008), considering that hepcidin levels are increased in iron overproduction (Ganz et al., 2006) this would suggest a therapeutic potential of HDAC inhibition in Hepatitis C through the reduction of iron toxicity .
9.3 Idiopathic pulmonary fibrosis (IPF)
Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal disease of unknown reasons characterized by acute lung injury. In the past it has been termed as cryptogenic fibrosing alveolitis (CFA). The disease is characterized by fibroblast proliferation and extracellular matrix accumulation (Fellrath et al., 2003).
Incidences and cases of IPF have been reported worldwide, to which the age range is 50-70 years old, the gender ratio is 1:1 or 2:1 representing of male:female with smoking as a main factor (Fellrath et al., 2003). In America clinical studies of IPF patients were analysed in rates of incidence and prevalence done between the period of January 1996 through December 2000. From the results, the prevalence rate was estimated to be a range from 4.0 per 100,000 persons aged 18 to 34 yr to 227.2 per 100,000 population among those 75 yr or older. The annual incidence was also estimated at a range from 1.2 to 76.4 per 100,000 (Fernandez Perez et al., 2010).
Diagnosing the disease can be done through 2 methods of analysis one is the sampling of lung biopsy to measure biochemical and pathological factors, the second is the non sampling of lung biopsy to assess physical characteristics of the patients. Overall several features need to be detected to identify the disease such as abnormal pulmonary function results such as evidence of restriction in FEV1 or impaired gas exchange, an age range of patient being over 50 and duration of the illness being more than 3 months (2000).
The inflammatory pathway of pulmonary fibrosis is complex especially if the cause or trigger is unknown. In the general pathology of pulmonary fibrosis there is a suggestions of huge amount of mediators, cytokines and cells are involved in the disease such as IL-1, TNF, Monocyte chemoattractant protein (MCP)-1, INF-Î³, IL-13, endothelia metalloproteinase 7 and most important fibroblast cells (Strieter, 2001).
A number of studies have identified the importance of HDAC involvement in the pathology and involvement in the development of IPF. Indicating the potential and need of developing of HDAC therapy for treatment of IPF. Administration of HDAC inhibitors SAHA and LBH589 (panobinostat) increased expression of COX-2 mediator PGE2 which is known to inhibit proliferation of fibroblast (Coward et al., 2009). Another study indicated the importance of HDAC inhibition in IPF therapy. Administration of HDAC inhibitor TSA in normal human lung fibroblasts (NHLFs) demonstration an inhibition of TGF-beta1-mediated alpha-smooth muscle actin (alpha-SMA) and alpha1 type I collagen mRNA induction both important mediators in fibroblast proliferation and function (Guo et al., 2009).
But in contrast administration of HDAC inhibitor LBH589 (panobinostat) increases expression of CXCL10 or Interferon gamma-induced protein 10 kDa (IP-10) in fibroblasts of IPF patients,IP-10 is known be a strong inducer of fibroblasts, suggesting HDAC enhancement is needed to inhibit fibroblast induction (Coward et al., 2010).
From the evidence and theory of HDAC involvement in inflammatory disease discussed. It is certain that HDAC is an important target for a variety of inflammatory diseases. However there is a problem there is not enough interest in this field.
The majority of the HDAC target therapy in various inflammatory diseases has been conducted on in vitro or in vitro conditions. Not much is known about HDAC in clinical trials of sepsis, rheumatoid arthritis, multiple sclerosis, Huntingdon disease,traumatic brain injury, haemorrhagic shock, stroke, hepatitis C and idiopathic pulmonary disease .
Most of the HDAC therapy in experimental and clinical trials has been targeted in inhibition in cancer clinical trials. The inflammatory diseases that HDAC targeted therapy in clinical trials are COPD and ulcerative colitis, in which HDAC enhancement has shown beneficial properties.
There might be a reason behind a lack of HDAC targeted therapy of inflammatory diseases. The first reason is the genetic involvement of HDAC in patients. The genetic composition differs from one person to another, targeting of HDAC can affect a different range of genes in one person compared to another person. Bringing different responses in the inflammation pathway of the disease.
The second reason is simply the lack of research and studies on the use of use of HDAC. As mentioned HDAC targeted therapy is more focused on cancer since it is a more dominant disease than other inflammatory diseases. Finally there is existence of alternative therapies for these anti-inflammatory diseases and entrusting a large amount of investment of HDAC research would be a huge risk.
But this wouldn't mean HDAC is completely expelled from the potential therapies of inflammatory diseases, the problem is there isn't enough attention on the subject. Hopefully in the future there will be more studies of HDAC therapy in inflammatory disease.