Influenza virus is one of the deadliest viruses in the world, capable of causing major viral outbreaks. This group of viruses belongs to the orthomyxovirus family. They are pleomorphic i.e. can take various forms. There are three types of the influenza virus, virus A,B and C. Strains A and B are responsible for the major influenza outbreaks and symptoms. Strain C causes minor symptoms. Influenza virus is responsible for a virulent form of flu. A lot of research has gone into identifying and generating therapies for dealing with influenza outbreak. Vaccines and antiviral therapy including drugs against influenza is under work. Vaccines would be ideally good for this purpose but there are concerns about the side effects associated with use of vaccines including allergic reactions, immune response to vaccines and chances of developing a rare syndrome. In this circumstance, anti-influenza drugs have gained a lot of importance.
Medicines developed to tackle influenza either directly targets the drug or the symptoms associated with the infection. The main class of drugs used currently target either neuraminidase or the viral protein but the challenge with development of drugs is development of resistance against the drug by the rapidly mutating and evolving viral genome. In addition, what makes it more difficult is the generation of new influenza strains in different species, which are capable of cross infecting spreading. This entails development of multi prolonged strategies aimed at combating influenza.
In this literature review, we look at the latest approaches and discoveries till now in the development of drugs against influenza. We are using a combination of internet based research along with books in this process. We will be trying to identify all the major class of drugs, their structures and efficacy and their mode of action and where possible, results of studies with them.
Viruses can be defined as acellular organisms which need a host cell for its survival and replication and consists of a nucleic acid encapsulated by a protein coat, which may or may not be enveloped in a membrane (Mcb.uct.ac.za, 2014) Once inside a host cell, they take over the host cell machinery and directs it into production of more viruses which ultimately leads to the death of the cell, when the viruses rupture the cell to be freed. Viruses are of different types based on the kind of nucleic acid they possess as genetic material, the kind of structure of their protein coats.
Influenza is one of the world’s most feared diseases on account of its ability to spread rapidly and cause epidemics. It brings about ‘flu” which leads to infection of the nose, throat, lungs and bronchi. The influenza virus belongs to the family orthomyxiviridae. The more prevailing influenza is called seasonal influenza and is caused by any of three known strains A, B and C. There are different subtypes of these strains. A and B viruses are the dominant ones. Of this the H1N1 and H3N2 subtypes of Influenza A is the most prevailing versions. (Anonymous, 2014)
The structure of the influenza virus is shown below . Existing and future drugs are based on the structure and life cycle of the virus. : (Racaniello, 2014)
Structure and Mechanism of infection: (Ruigrok, 1998)(Colman, 1998) (Varghese 1998) (Steinhauer, 1998)
The virus is typically spherical and is an enveloped virus. It consists of the following parts:
- Envelope –outer, lipid bilayer and created from the host cell membrane of the cell in which the virus is growing. The envelope has characteristic spike like projections embedded in it which protrude out. These are the membrane glycoproteins HA and NA. HA is haemagglutinin and NA is Neuraminidase. The HA consists of three monomers while the NA is a tetramer (Stainhauer 1998, Colman 1998). HA is the major envelope protein and its 3D structure has been determined. The HA is synthesized as a single polypeptide unit which is subsequently divided into its two sub units HA1 and HA2. This split is necessary for the fusion between viral cell envelope and the host cell membrane (Lam and Krug, 2001) The HA and NA proteins are used to classify the subtype of the virus. Besides these other proteins called M1 and M2 are present. M1 is a matrix protein and M2 an ion channel. M1 confers rigidity and strength to the virus particle while M2 is an ion channel, which regulates the pH of virions. Both the NA and M2 proteins are subject of targeted antiviral strategies. HA is the primary instigator of antibody responses by the host. The Neuraminidase or Sialidaseconsists of four subunits and is an exoglycohydrolase which cleaves the thesialic acid N-acetylneuraminic acid residues linked to glycoconjugates in the lungs and respiratory tract thus enabling movement of the virus in the respiratory tract (Colman and Ward, 1985; Palese et al 1974, Liu et al 1995, Ruigrok, 1998; Colman, 1998) (Varghese 1998) (Steinhauer, 1998)
- Viral genome – RNA is the genetic material and there are 8 RNAs for influenza A. Each codes for viral proteins. The uniqueness of influenza virus is that its genome is segmented. EachRNA is complexed with proteins (polymerases and nucleoproteins) to form a RNA segment.
We will be examining available literature sources on the methods of treatment for influenza, especially pertaining to synthesis of drugs. The sources we used for our literature search are Pubmed (www.ncbi.nm.nih.gov) which is the most widely known resource site for journals and information related to biological resources. Also, Google search was used to independently ferret out independent papers and Wikipedia to obtain some papers regarding this topic.
Literature review has revealed that the kind of treatments available for Influenza belongs to two classes: Vaccines and Antiviral treatments. While vaccines have been in use for sometime and still remain popular for treatment in the young and elderly, attempts to generate antiviral drugs have also been in the process, the reason being that during times of epidemics, it would be more useful to have drugs on hand rather than wait for the synthesis of vaccines. Research into drugs targeting influenza virus have had some successes and some challenges. (Stiver, 2003)
Traditionally, the two major classes of drugs which have been in use are the adamantanes (amantadine and remantadine), and 2) inhibitors of influenza neuraminidase (oseltamivir and zanamivir). These drugs target different aspects of the viral life cycle including the viral coat formation, Nucleotide synthesis and transcription. But some new drugs differing from the traditional class are in progress.
- Traditional Influenza drugs
Adamantanes- This includes amantadine and remantadine. Amantadine is 1-adamantylamine or 1-aminoadamantane and has been used both as an antiviral as well as a treatment for Parkinson disease. Rimantadine is similar to adamantine. As the viral has developed resistance to this drug, since 2009 it is not recommended by governments to take this drug. In addition, the drug also has several side effects on the gastro intestinal and nervous system.
Mechanism of action: it acts by interfering with the operation of the M2 ion channel viral protein. This does not allow the virus to get uncoated and thus prevents its spreading. This drug is effective only with influenza a.
- Neuraminidase inhibitors
Neuraminidase is the enzyme which cleaves sialic acid residues present on the cellular receptors which are responsible for attaching the virus particles to the cell. Inhibitors to this enzyme compete with the substrate for the enzyme’s active site, thus interfering in the function of the enzyme i.e. preventing the removal of sialic acid residues necessary for the attachment of the virions to the host cell. As a result, the infection is stopped. The virus particles cannot spread further. The two main drugs which are very popular neuraminidase inhibitors are Oseltamivir and Zanamivir. These are second generation drugs. These drugs are widely used as they inhibit all classes of influenza whereas the M2 inhibitors function only against influenza A. (Stiver, 2008)
Oseltamivir is also known as tamiflu. It is an orally taken inactive drug which becomes active when ingested. The problems with oseltamivir usage include side effects and drug resistance. In fact between 2007 and 2009, there was high resistance against this drug by the virus.
Zanamivir is another neuraminidase inhibitor. Another neruraminidase inhibitor is Laninamiviroctanoat. On inhalation, it is converted into its active form, laninamivir, which persists in the lungs for a long time and is enough to treat influenza. It has also demonstrated activity against oseltamivir resistant drugs. (Tappenden et al 2010)
Another neuraminidase inhibitor is Laninamiviroctanoate. On inhalation, it is converted into its active form, laninamivir, which persists in the lungs for a long time and is enough to treat influenza. It has also demonstrated activity against oseltamivir resistant drugs. (Ikematsu and Kawai, 2011)
- Ribavirin – A third class of drugs are those which interfere with viral replication. Ribavirin is a ribonucleoside analogue. It has been found to interfere with viral mRNA synthesis and mRNA capping by competing with the ribonucleosideguanosine. It reduces the size of the cellular pool of Guanosine 5’ -triphosphate. Besides influenza, it is also shown to be beneficial for combatting various viral fevers which cause haemorrhage as well as hepatitis C. (Crotty et al 2002, Carter and Saunders, 2007)
Its carboxamide structure makes it resemble adenosine or guanosine, the bases which are required for RNA synthesis. It thus acts as a base analog and incorporation of ribavirin molecules, instead of the normal bases, would create mutations in the newly synthesised RNA molecules which may be detrimental to the viruses (Crotty et al 2002). What is extremely curious and interesting about ribavirin is that it can also inhibit the activity of DNA viruses, even though its structural specificity is for ribonucleosides. Its most common derivative is taribavirin. It has been used against influenza A but is not considered a very effective drug against it. ( Uchide et al, 2010) (Wray et al 1985, Wong and Yuen, 2006)
Burch and colleagues in 2009 conducted a study in which Amantidine was not included as there was no new information available on its action. The studies showed that the antiviral drugs which target theneuraminidase enzyme of the virus were effective in reducing the length of time for the symptoms to remain. They reduced by 0.5-1.5 days average. These drugs included oseltamivir and zanamivir. The impact on effects was more observable in at risk groups than in healthy adults and points towards the fact that these new drugs may be used with greater effect in at risk population (Burch et al 2009) Another study by Tappenden and group also in 2010 highlighted the limited evidence about the success of amantidine in preventing seasonal confirmed influenza cases. Whereas, evidence for the success of OSeltamivir and Zanamivir in preventing Seasonal Lavoratory confirmed Influenza was confrmed. (Tappenden 2009) (Cdc.gov, 2014)
New solutions to influenza
Because of the challenges offered by the influenza virus against drugs such as display of resistance, new and alternative drugs targeting new routes to combating influenza are under action.
- Cyclosporin A is an immunosuppressant (Laupasis 1982) which has been used in transplantation cases to reduce the rejection by the immune system of organ transplants. Kaminsky( 2008) mentions that it has unilateral influence on the activity of the T cells
Structure: cyclic polypeptide consisting of 11 amino acids, produced naturally as a metabolite by certain fungi It is: [R-[R*,R*-(E)]]-cyclic(L-alanyl-D-alanyl-N-methyl-L-leucyl-N-methyl-L-leucyl-N-methyl-L-valyl-3-hydroxy-N,4-dimethyl-L-2-amino-6-octenoyl-L-α-amino-butyryl-N-methylglycyl-N-methyl-L-leucyl-L-valyl-N-methyl-L-leucyl).
Mechanism of action
Role against influenza
SO far widely used in immunosuppression strategies, a recent paper by Hamamoto et al have highlighted how it can be used to tackle influenza virus action. They found that CsA could suppress the growth of influenza virus is A549 cells by interfering with a stage in the life cycle. The step is believed to be post viral protein synthesis possible assembly and generation of new virus particles.
- Nucleozin1: Nucleozin is the type of a new class of antiviral drugs which are being tried out to combat influenza. These drugs are believed to act on the Influenza A virus replication by interfering with the transport of the viral gene segments in the cytoplasm.
Structure:(Amorim et al 2013)
Mechanism of action
Nucleozin and its derivatives were thought to act as inhibitors of viral nucleoprotein.”These inhibitors are thought to act as “molecular staples” that stabilize interactions between NP monomers, promoting the formation of nonfunctional aggregates “( Amorim et al 2013)
The scientists have shown that this drug has effects at both initial and final stages of the viral life cycle. Inhibitorsof nucleoprotein not only inhibit viral RNA replication and synthesis at the start of the viral life cycle, but also block the intracellular transport of the ribonucleoproiteins in association with rab11, causing their aggregation into clumps and this resulted in smaller number of viral particles. This effect is mediated through their action on the viral ribinucleoprotein (Amorimetal 2013) Thus,nucleozin has emerged as a novel antiviral drug which operates through a distinct mechanism
Further, Cheng and colleagues in 2012 have found that certain Nucleozin related derivatives (compound 3b) synthesized from Nucleozin 1 used as lead, inhibits replication of different strains of H3N2 and H1N1 as well as amantidine and oseltamivir resistant viruses. This is a great break in the development of antiviral drugs.
Several compounds have also been identified in large scale drug screening procedures, many of which display inhibition of rNA directed RNA polymerase activity. (Cy, 2010) These included nucleozin and its analogs. Other compounds such as ant oxidants have also been considered as potential antiviral drugs e.g. Superoxide dismutases conjugated with polymers, ribavirin etc, thiol antioxidants such as pyrrolidinedithiocarbamate(Uchide et al 2010)
What makes developing lasting cures for influenza extremely challenging are the following reasons:
- The RNA genome can mutate rapidly as a result the kind of proteins and enzymes encoded by it will keep changing and this makes devising permanent targeted therapies difficult. This helps increase horizontal spread of the virus
- Its multiple genomic segments can assort and reassert themselves during co-infection thus generating a far greater diversity of strains
This leads to the creation of new types of antigens and developing antibodies becomes a tedious and challenging task.
Because of this, the drugs developed against the virus can lose its efficacy and it may develop antiviral drug resistance of for instance, of some old drugs which are not recommended in against present-day infections of influenza include Amantidine, Tamiflu® and Relenza®. These drugs cause side effects as well as are already resisted by the new set of viruses. (Barik, 2012, p. 104)
All of the information for this literature reviews was obtained through searches of literature on the internet directly through Google or through the Pubmed website (www.ncbi.nlm.nih.gov) as well as through Wikipedia for searching relevant journals. The terms used for starting off the literature searches were as follows: Influenza and anti-virus drugs, antiviral drugs and H1N1, influenza virus and structure, influenza virus and life cycle. It resulted in the retrieval of sites providing general information on influenza such as that published by health organization giving information on its treatment, patient care, statistics.
Example of such results was:
- Using Google
There are strengths and weaknesses using Google as a search tool for literature review. Google is fast, powerful and brings up results in a jiffy but there may be a lot of unwanted information which you have to filter out. So you need to use effective key words. Many times, I needed to scroll down a lot to get some relevant information or papers or websites. Some of the sites were then used as sources to other sites. For example the website shown (http://www.uptodate.com/contents/pharmacology-of-antiviral-drugs-for-influenza) has references to other papers on influenza. So the search can be linked in such ways.
- Using Pubmed
Searches resulted in many papers on influenza. A few were useful and provided information relevant to this review. These include papers by Amorim et al (Nucleozin), Stiver (2003), Barik (2012), Colman etc. Others were far more into specifics such as creating high throughput derivatives and mechanisms of actions, as well as relating to treatment, which were beyond the scope of this review.
- Using Wikipedia
This also provided a lot of information and especially literature sources, which could be used for the research required for this study. Understanding the structure of the influenza virus and the molecular biology of its life cycle is crucial to the developing of highly specific antiviral drugs against it. There has been in-depth research carried out, due to which we know quite well the structure-function aspects of the influenza virus, its proteins, and its genome. The more specific details have resulted in the generation of drugs which target the virus at different points and as the literature review shows there is plenty of work being carried out on the development of new drugs such as nucloezin based derivatives which will be able to overcome the weaknesses of the older ones such as amantidine.
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