key: cord-0006271-qeq33nns
authors: McKimm-Breschkin, Jennifer L.
title: Management of Influenza Virus Infections with Neuraminidase Inhibitors: Detection, Incidence, and Implications of Drug Resistance
date: 2012-08-23
journal: Treat Respir Med
DOI: 10.2165/00151829-200504020-00004
sha: 00ae60473d68cce817c03b7e329e0546d99736f2
doc_id: 6271
cord_uid: qeq33nns

Although influenza vaccination remains the primary method for the prevention of influenza, efficacy may be limited by a poor match between the vaccine and circulating strains and the poor response of elderly patients. Hence, there is an important role for antiviral therapy in the management of influenza. While amantadine and rimantadine have been available for the treatment of influenza in some countries for several years, they are only effective against influenza A viruses, they can have neurological and gastrointestinal adverse effects, and resistant virus is rapidly generated. Neuraminidase inhibitors, a new class of drug, are potent and specific inhibitors of all strains of influenza virus, and they have minimal adverse effects. The greatest benefit is seen in those patients presenting <30 hours after development of influenza symptoms, those with severe symptoms or those in high-risk groups. In addition to treatment of the infection, both drugs are effective prophylactically and have been shown to limit spread of infection in close communities, such as families and in nursing homes. No resistant virus strains have been isolated from normal individuals treated with zanamivir. Resistant virus can be isolated from approximately 1% of adults and 5% of paediatric patients with influenza treated with oseltamivir. However, infectivity of mutant viruses is generally compromised. Governments spend millions of dollars on influenza vaccination campaigns; however, once influenza virus is circulating in the community, vaccination cannot limit the spread of disease. A greater promotion of the use of neuraminidase inhibitors for the treatment and prevention of influenza could have a significant impact on limiting its spread. This could result in saving millions of dollars, not only in direct costs associated with medical and hospital care, but also significant savings in indirect costs associated with the loss of productivity at work, school and home environments. For the benefit of all communities, there needs to be a greater awareness of the symptoms of influenza and the efficacy of neuraminidase inhibitors in disease treatment.

Although influenza vaccination remains the primary method for the prevention of influenza, efficacy may be Abstract limited by a poor match between the vaccine and circulating strains and the poor response of elderly patients. Hence, there is an important role for antiviral therapy in the management of influenza. While amantadine and rimantadine have been available for the treatment of influenza in some countries for several years, they are only effective against influenza A viruses, they can have neurological and gastrointestinal adverse effects, and resistant virus is rapidly generated. Neuraminidase inhibitors, a new class of drug, are potent and specific inhibitors of all strains of influenza virus, and they have minimal adverse effects. The greatest benefit is seen in those patients presenting <30 hours after development of influenza symptoms, those with severe symptoms or those in high-risk groups. In addition to treatment of the infection, both drugs are effective prophylactically and have been shown to limit spread of infection in close communities, such as families and in nursing homes. No resistant virus strains have been isolated from normal individuals treated with zanamivir. Resistant virus can be isolated from approximately 1% of adults and 5% of paediatric patients with influenza treated with oseltamivir. However, infectivity of mutant viruses is generally compromised. Governments spend millions of dollars on influenza vaccination campaigns; however, once influenza virus is circulating in the community, vaccination cannot limit the spread of disease. A greater promotion of the use of neuraminidase inhibitors for the treatment and prevention of influenza could have a significant impact on limiting its spread. This could result in saving millions of dollars, not only in direct costs associated with medical and hospital care, but also significant savings in indirect costs associated with the loss of productivity at work, school and home environments. For the benefit of all communities, there needs to be a greater awareness of the symptoms of influenza and the efficacy of neuraminidase inhibitors in disease treatment.

vaccination is not effective at the point of the outbreak, since it may take 2-4 weeks to achieve protective levels of immunity after vaccination. Hence, there is an important role for antiviral therapy Influenza is a severe infection of the upper respiratory tract in the management of influenza. responsible for significant morbidity and mortality throughout the world. It is estimated that in the US alone approximately 300 000 2. Anti-Influenza Therapy hospitalisations per year are attributed to influenza with up to 35 000 deaths. [1] Other viruses including parainfluenza virus, re-There are two classes of drugs available to control the spread of spiratory syncytial virus and Coxsackie viruses can all cause acute influenza, each targeting different virus proteins. The first class, respiratory infections. Although there are several rapid diagnostic the adamantanes, blocks the ion-channel function of the M2 protetests for influenza, most are complex and have low sensitivities in of the influenza virus. The second group, neuraminidase inhibi-(reviewed in Nicholson et al. [2] ). However, onset of influenza tors, targets the neuraminidase of the virion. [9] illness is usually rapid, and when influenza virus is known to be In order to understand how anti-influenza drugs work, it is circulating in the community, clinical diagnosis, based on fever necessary to understand how the influenza virus replicates. There plus two other symptoms including cough, sore throat, headache are two types of influenza viruses that cause severe disease in or myalgia, has been shown to be accurate in approximately 70% humans, influenza A and B. Virions have two glycoprotein spikes of cases. Diagnosis based on fever plus cough was shown to be on their surface, the hemagglutinin and the neuraminidase, which accurate in up to 79% of patients in one study; however, a second allow the virus to enter and leave cells. Hemagglutinin binds to recent study reported a predictive value of only 40-60% for these cellular receptors that contain terminal sialic acids. Once bound to two symptoms. [3] the receptor, the virus is endocytosed and transported into the Although most symptoms of influenza are common to all ages, cytoplasm. The virus M2 protein acts as an ion channel allowing children tend to have higher fever than adults, as well as drowsithe penetration of hydrogen ions from the endosome into the virus. ness and gastrointestinal symptoms, including abdominal pain,

The low pH leads to hemagglutinin unfolding and fusing with the diarrhoea and vomiting. Elderly patients tend to have a higher endosomal membrane, releasing virus nucleic acid into the cell. incidence of lower respiratory tract symptoms, including sputum Replication of the viral RNA and production of new virus proteins production, wheezing and chest pain. [4] Complications generally then occurs. Newly synthesized virions assemble and bud at the involve the respiratory tract, with acute bronchitis and pneumonia cell membrane. Since new virions all contain hemagglutinin, they being the most frequent, [5] but young children and infants are at will bind to the cellular receptors and to sugars on other virions as risk of acute otitis media, bronchiolitis and croup. [6] While the they bud from the cell. The second virus surface protein, direct costs of primary medical care and hospitalisations run into neuraminidase, is an enzyme that removes sialic acids from the billions worldwide, there is an even greater cost to the community cellular receptors. [10] Since hemagglutinin can no longer bind to with loss of productivity either in the workplace or home environthe cell, this allows release of progeny virions. In addition, removments.

al of sialic acid from virus proteins prevents newly formed virions Vaccination remains the primary method for prevention of from binding to each other and aggregating. Neuraminidase may influenza and its potential complications. However, vaccination also play a role in facilitating movement of the virus through the has its limitations. Influenza viruses undergo continual mutation, mucus layer in the respiratory tract. [11] so that virus strains selected for the vaccine may be a poor match to the currently circulating strains; therefore, immunity may not be 2.1 Adamantanes protective. In healthy individuals, the protective efficacy is estimated to vary between 70% and 90%, [7] whereas efficacy in the The adamantanes block the function of the M2 protein at low elderly or immunosuppressed can be as low as 40-60%. [8] In concentrations, and at very high concentrations act indirectly on addition, when a severely ill patient presents in the clinician's hemagglutinin, preventing fusion of the virus and cell memoffice with suspected influenza or in the event of a pandemic, branes. [12] Amantadine has been available in the US for prophylax-is and treatment of influenza since 1976 and rimantadine has been There have been recent reports of enhanced potency of multivalent zanamivir by Sankyo in Japan [23, 24] and Biota in Ausavailable since 1993. However, these drugs have not been aptralia, [25] where a single dose of drug may be sufficient for protecproved for use in all countries. Although these drugs may be the tion. cheapest option for treating influenza, they also have a number of limitations in their use. They are only effective against influenza A 3. Clinical Trials viruses, since influenza B viruses do not have an M2 protein. In addition, 10-30% of patients suffer from intestinal and CNS adverse effects; this can exacerbate problems in the elderly, who 3.1 Treatment may already be disoriented. [13] Furthermore, resistant virus is readily generated in up to 30% of patients. [14] Mutant viruses are

still virulent and readily transmitted, [15, 16] and can be isolated from Randomized, double-blind, placebo-controlled trials have been patients in the community who have never been exposed to these undertaken for both zanamivir and oseltamivir. A number of inhibitors. [17, 18] articles have reviewed and summarized data from the clinical trials involving both drugs. [26] [27] [28] [29] [30] The primary clinical endpoint was the length of time to alleviation of symptoms (fever, myalgia, head-

ache, sore throat or cough) for at least 24 hours. Reduced virus shedding was also observed with both drugs. However, it has Neuraminidase inhibitors are highly potent and specific inhibirecently been reported that more than 10 3 infectious units/mL of tors of both influenza A and B viruses. They were designed using a influenza virus could still be isolated from some children treated new approach to drug development based on understanding the with oseltamivir, even after 5 days of treatment.

[31] structure of the target molecule and how it interacts with its For zanamivir, a mean 1-1.5 days of earlier symptom relief was substrate. The determination of the 3-dimensional structure of the observed for all patients, including adults and paediatric patients influenza neuraminidase in complex with sialic acid led to the compared with inhaled placebo [32] . Despite a concomitant reducdesign of zanamivir. [9] Zanamivir interacts with a group of amino tion in the severity of symptoms, the perception among many acids in the active site of neuraminidase, which are conserved in clinicians and public is that 1-day of symptom improvement is not all influenza A and B strains. Zanamivir blocks the action of significant. However, when the data was analysed among different neuraminidase, which prevents the cleavage of sialic acid on the risk groups, more significant results were demonstrated. Those cell receptors, thus preventing release and spread of the newly that derived the greatest benefits (2-3 days of symptom relief) formed virions. Since neuraminidase activity is also thought to were high-risk patients, those presenting with severe influenzafacilitate the passage of virus through mucus, blocking related symptoms or those who had symptoms for <30 hours. neuraminidase activity may also prevent the virus from reaching Older patients, aged >50 years, presenting with severe symptoms respiratory cells. Zanamivir is administered by oral inhalation, derived up to 7 days of benefit, as in this group, untreated patients achieving high concentrations of drug in the respiratory tract, the experienced up to 11.5 days of symptoms. [32] site of virus replication at a dosage of two inhalations of 5mg each

In addition to earlier symptom relief, patients returned to nor-(total dose 10mg) twice daily for 5 days. Based on the efficacy of mal activities sooner, and there was a significant decrease in the zanamivir, a second inhibitor, oseltamivir, was subsequently deuse of symptom relief medication and antibacterials. Adverse veloped. [19] Oseltamivir is taken orally as the ethyl ester prodrug, effects were no different between the placebo-and zanamiviroseltamivir phosphate, which is converted by hepatic esterases treated groups. Since zanamivir has low systemic availability, no into the active form, oseltamivir carboxylate. The drug is adminisdosage adjustment is necessary for patients with impaired renal tered at a dosage of 75mg twice daily for 5 days.

function. While there were reports of bronchospasm post-release, Two other neuraminidase inhibitors have been developed; howtrials specifically targeting patients with mild-to-moderate asthma ever, neither is being pursued. Peramvir (BCX 1812, RWJ or COPD showed that zanamivir was an effective treatment and 270201) originated by Biocryst Pharmaceuticals [20] was an effechad a similar safety profile to the inhaled lactose placebo. [33] There tive inhibitor in vitro; however, significant improvements in pawere no adverse effect on pulmonary function; however, the pack tients' outcomes were not observed in clinical trials. Abbott was insert recommends that patients should have a fast-acting also developing a neuraminidase inhibitor (A 315675) [21] which bronchodilator available. Interestingly, there were also five cases displayed in vitro inhibitory activity. [22] However, this drug has not of bronchospasm reported in the oseltamivir clinical trials: [34] one been taken into clinical trials.

in the placebo group and four in the oseltamivir group, suggesting that bronchospasm may be a symptom associated with influenza ing receptors for pneumococcal adherence. Oseltamivir improved infection.

survival of mice, independent of viral replication and morbidity from influenza. [42] Thus, treatment of influenza with Post-marketing surveillance of more than 15 000 patients neuraminidase inhibitors may play an important role in preventing showed that symptoms improved within 2 days of initiation of exacerbation of pneumonia. treatment with zanamivir in >70% of patients and two-thirds of patients returned to normal activities within 3 days. [35] Despite the

low incidence of zanamivir prescribing by clinicians, due to the Neuraminidase inhibitors are effective against all subtypes of perception that the inhaler is difficult to use, particularly by the neuraminidase, which is of critical importance in treating any elderly, [36] post-marketing surveillance showed that 89% of papotential pandemic strain of influenza virus. Testing of approxitients aged >65 years were comfortable with using the inhalmately 1000 human H1N1, H3N2, H1N2 and influenza B viruses, er. [37, 38] has demonstrated that zanamivir is slightly more effective against N1 and influenza B viruses, and oseltamivir is slightly more

effective against N2 viruses. [43] In addition, neuraminidase inhibi-For oseltamivir, a benefit of 1-1.5 days of earlier relief of tors have been shown to be specifically effective against symptoms was observed for all patients, and a greater benefit of 2 neuraminidase from the virus responsible for the 1918 pandemdays was observed for those presenting at less than 24 hours after ic, [44] as well as against the H5N1 and H9N2 avian virus strains onset of symptoms. [26] [27] [28] 30] Patients in the oseltamivir treatment responsible for the 1997-1999 outbreaks of avian influenza in group returned to normal activities sooner, and there was less use Hong Kong. [45, 46] The highly pathogenic avian H5N1 viruses, of antibacterials and relief medication compared with the placebo which were circulating in Asia during the period 2003-4 were also group. Likewise, children treated with oseltamivir also expersensitive to neuraminidase inhibitors, [47] but interestingly were ienced a 1.5-day benefit of symptom relief, returned to normal resistant to the adamantanes. [48] There is a perception that multiple activities sooner, had a reduction in prescription of antibacterials organ failure observed in the few humans who have died as a result and fewer complications, including otitis media, [39] than the placeof an H5N1 infection, is due to systemic spread of virus and bo control group. Separate data on the efficacy of oseltamivir multiplication in the peripheral organs. Hence, only a drug that treatment of high-risk patients have not yet been published. Adacts systemically would be effective. However, post-mortem exverse effects included nausea and vomiting, with 17% experiencaminations of humans infected with the chicken virus strains have ing transient nausea in the oseltamivir group compared with 7% in failed to find virus anywhere other than in the respiratory tract and the placebo group. [40] Effects were primarily in the first 48 hours of lungs. [49] Similarly, animal models of chimpanzees infected with treatment and could be partially reduced by taking the drug with the avian influenza viruses show that although the animals die of food. A study in experimental influenza found that administration organ failure, virus is only detected in the respiratory tract and of oseltamivir with food reduced the incidence of gastrointestinal lungs. [50] Destruction of other organs appears to be due to high disturbances from 31% to 7%. [40] Dosage adjustment to 75 mg/day levels of cytokines released in response to influenza virus infecis needed for patients with creatinine clearance <30 mL/min. For tion. Since the infection is still spread by the respiratory route, and patients with clearance rates <10 mL/min, caution is advised even limited to the respiratory tract, inhaled zanamivir should be just as with the lower dose of oseltamivir (package insert). There has also effective as oral oseltamivir. been a recent report of death in juvenile rats treated with high doses of oseltamivir. [41] Brain levels of oseltamivir were 1500-fold 3.2 Prophylaxis higher than those of adult animals exposed to the same dose. It is hypothesized that immaturity of the blood-brain barrier caused the In a randomized double-blind, placebo-controlled trial, particitoxicity. The US FDA has therefore instructed Roche to include a pants began 4 weeks of zanamivir at the start of an influenza A warning in the package insert, indicating that oseltamivir should outbreak. Zanamivir was 84% effective in preventing laboratory not be administered to children <1 year of age. Oseltamivir is confirmed illness with fever. [51] Interestingly, many patients were administered as syrup to paediatric patients.

apparently infected asymptomatically since many developed an Co-infection with Streptococcus pneumoniae and influenza can immune response without associated symptoms, thus providing have a synergistic effect on disease severity, leading to excess protection from reinfection. In a study evaluating prevention of mortality. [42] In a mouse model, it was demonstrated that the influenza in families, where both index cases and family contacts stripping of sialic acid from the lung by influenza virus were treated, zanamivir was 79% effective in preventing influenza neuraminidase potentiated development of pneumonia by expos-in contacts. Zanamivir was effective against both influenza A and B, and the index cases had symptoms for 2.5 days less than

placebo-treated controls. [52] In a second trial where the index case As discussed above, the influenza M2 inhibitors have a very received relief medication only, zanamivir treatment of household poor resistance profile, rapidly generating resistant viruses, which contacts had an 81% protective efficacy against laboratory-conare virulent and transmissible. Furthermore, resistant virus can be firmed influenza, and protective efficacy was similarly high isolated in untreated patients in the community. Because against both influenza A and B. [53] Zanamivir was also effective in neuraminidase inhibitors are a new class of drug, there are cona nursing home outbreak, where despite a 90% vaccination rate cerns about the potential for the emergence and spread of resistant there was an outbreak of A/Sydney/05/97 H3N2-like virus, a variants. The regulatory authorities have requested surveillance be component of the vaccine. Amantadine therapy did not control the carried out to monitor both the potential drift in the baseline spread, and amantadine-resistant viruses were isolated. In the 2 sensitivity of isolates and the emergence of resistance in individual weeks after zanamivir prophylaxis, no new cases of influenza were isolates. The global neuraminidase Inhibitor Susceptibility Netconfirmed. [54] work (NISN) [61] was established to coordinate testing of clinical Prophylaxis studies with oseltamivir in patients challenged isolates to determine baseline sensitivities of isolates prior to introduction of the inhibitors into clinical practice, as well as postwith influenza A virus showed that oseltamivir was 61% effective release. Of approximately 1000 isolates tested, collected prior to in preventing influenza infection based on serology, and no patient drug release based on sensitivities of their neuraminidases, there shed virus. [40] Oseltamivir did not reduce infection rates in the was no evidence of naturally occurring resistance to either drug in influenza B virus challenge, although it did reduce the virus titre any of the isolates. [43] It is proposed that post-release monitoring and duration of shedding. [55] A further trial where patients were should be carried out for 5 years. treated for 6 weeks during a local influenza outbreak demonstrated a protective efficacy of 74%. [56] In a household contacts trial where 4.2 Resistance due to Hemagglutinin Mutations the index case was untreated, the overall protective efficacy against clinical influenza was 89% for individuals and 84% for Although hemagglutinin mutations are the predominant mutahouseholds. [57] In what was thought to be an influenza B virus tion in cell culture, their role in altered sensitivity of clinical outbreak in a nursing home, oseltamivir prophylaxis reduced the isolates is not known. Mutations in hemagglutinin are located in incidence of influenza infection by approximately 50%, compared and around the regions of the molecule involved in binding to the with a home where no treatment was used. [58] In a frail elderly cellular receptor. Reduced affinity of hemagglutinin for the sialic population, oseltamivir prophylaxis resulted in a 92% reduction in acid means the newly formed virions do not bind as strongly to the the incidence of laboratory-confirmed clinical influenza compared cell receptors, and they can therefore still elute even with reduced with placebo. [59] Consistent with the reduced efficacy of vaccinaneuraminidase activity. Thus, hemagglutinin mutations [60] result in decreased sensitivity to all neuraminidase inhibitors. Decreased tion in elderly patients, 12 of 13 patients who became infected in sensitivity due to hemagglutinin mutations is demonstrated by the the placebo group had been vaccinated. growth of virus in concentrations of either drug known to inhibit replication of wild type virus. However, the altered binding is 4. Drug Resistance specific to the type of cell and receptor. Hence, a human clinical isolate may not have altered sensitivity in the Madin Darby Canine Since neuraminidase inhibitors were designed at the molecular Kidney (MDCK) cells most commonly used to grow influenza in level to interact only with the active site of the virus the laboratory. One virus containing a hemagglutinin mutation at neuraminidase, mutations altering the ability of the drug to bind Arg198Thr (as well as an neuraminidase mutation; see section 4.3) are likely to have detrimental effects on the ability of the virus was isolated from an immunocompromised child treated with neuraminidase to function normally. Because of the differences in zanamivir. [62] However, the virus was sensitive to zanamivir in the the chemical structures of zanamivir and oseltamivir, they bind MDCK cells; hence, its role in in vivo resistance is not clear. An slightly differently in the enzyme active site. This means that hemagglutinin mutation, Ser262Asn was detected in a child in a mutations may affect the binding of one, but not necessarily both recent Japanese study, [31] in conjunction with a neuraminidase drugs. Decreased sensitivity to the inhibitors can be generated in mutation (Arg292Lys); however, its role in resistance has not yet cell culture in the laboratory, but only after extensive exposure of been confirmed. viruses to the drugs. [60] Mutations in vitro are found not only in Two H3N2 viruses with hemagglutinin mutations at Arg229 neuraminidase, as expected, but also in hemagglutinin.

were isolated from untreated patients. [63] Mutations at this residue in hemagglutinin conferred decreased sensitivity to both neuraminidase inhibitors in H1N9 viruses passaged in vitro. [64] Although both the H1N9 and H3N2 viruses had reduced sensitivity to neuraminidase inhibitors in MDCK cells, the H1N9 mutant viruses had reduced sensitivity in mice, [60] whereas the human H3N2 isolates were sensitive in ferrets, which are thought to have receptors more like those in humans. Hence, these mutations may have no effect on drug sensitivity in humans. Since the cells used in the laboratory have different types of receptors compared with human respiratory cells, there is currently no suitable cell for assessing the role of hemagglutinin in the development of resistance in human clinical isolates. One group is attempting to make an MDCK cell which has a higher density of human like sialic acids; [65] however, this has not yet been tested with a panel of known resistant hemagglutinin mutants.

Drug resistance due to mutations in neuraminidase are detected by screening viruses in an enzyme inhibition assay. To date, this is the only accepted method of determining drug resistance in clinical isolates. Neuraminidase mutations can render the enzyme from 10-to 10 000-fold less sensitive to the inhibitors. Mutations in neuraminidase can be both drug and virus neuraminidase subtype specific. [60] Both zanamivir and oseltamivir are based on the structure of a product of sialic acid generated during catalysis, 2,3-dehydro-2-deoxy-N-acetylneuraminic acid (DANA) [ figure  1 ]. However, zanamivir has only a single substitution of a guanidinium group at the 4′ position on the sugar ring. Oseltamivir has several differences, since it contains a cyclohexene ring instead of the sugar ring, and has two substitutions on this ring. It has a substitution of an amino group at the 4′ position and a large pentyl ether group replacing the glycerol side chain at the 6′ position. Binding of oseltamivir requires the movement of amino acids in the active site of neuraminidase to accommodate the larger side chain, whereas zanamivir does not. Because of the differences related as possible to the natural ligands of the target. This means in their chemical structures, there are specific mutations that affect that the greater the difference between the inhibitor and the natural the binding to each of the inhibitors. For example, mutations substrate, the greater the possibility that mutations can arise that preventing the movement of the amino acids in the active site affect binding of the inhibitor without affecting binding of the confer a higher level of resistance to oseltamivir, whereas those that affect the interaction with the guanidinium group confer natural substrate. [66] greater resistance to zanamivir. A virus with an N9 neuraminidase There has not yet been any documented case of clinical resishas been used as a model virus for resistance analysis in vitro, tance in a previously healthy individual treated with zanamivir. since the 3-dimensional structure of neuraminidase is known.

The only resistance seen after zanamivir treatment has been in an Analysis of how single mutations affect drug binding at the immunocompromised child after a prolonged infection with influstructural level has provided valuable insights into the mechaenza B and treatment with zanamivir for 15 days. [62] This virus had nisms of drug binding. It has also led to the hypothesis that a a mutation at Arg152Lys in neuraminidase, and Arg198Thr in general strategy for drug design when the target has a high mutation frequency is to design the inhibitor to be as closely hemagglutinin, neither of which have been seen in vitro. This virus is less sensitive to both zanamivir and oseltamivir in enzyme Since all mutations so far are located in or around the active site inhibition assays. [43] of the enzyme, the mutant neuraminidases have demonstrated either reduced stability, reduced enzyme activity, or reduced infec-Mutations in neuraminidase causing resistance to oseltamivir tivity in vitro or in animal models. [60, 73, 74] have arisen both in challenge studies and in patients with naturally

Poor infectivity of mutant viruses has been shown to affect acquired infections. Rates of resistance are estimated to be around virus transmission in animal models. [73] Ferrets infected with an 1% in the adult population and 5% in pediatric patients. [39, 67] There A/Sydney/5/97 (H3N2) clinical isolate carrying the Arg292Lys is, however, a recent report of resistant virus being isolated from 9 mutation did not transmit mutant virus to any contact animal. of 50 (18%) children treated with oseltamivir in a Japanese Unexpectedly, wild-type virus was isolated in contact animals; study. [31] In addition, wild-type virus was isolated from 24 patients apparently the mutant virus underwent reversion in the original despite several days of oseltamivir treatment.

infected animal. Thus, it could be theoretically possible that Mutations seem to be specific to the subtype of influenza virus.

oseltamivir treatment could generate a resistant mutant, which The most common mutation in N2 viruses is Arg292Lys, which may result in continued spread of a wild type revertant virus after gives 1000-to 10 000-fold resistance to oseltamivir in an enzyme cessation of treatment. sensitivity assay, and 10-to 100-fold reduced sensitivity to However, despite the viruses being compromised in their zanamivir. Although this mutation has been generated after pasgrowth in vitro, a more recent publication has demonstrated transsage of the N9 and N2 viruses in oseltamivir or zanamivir in cell mission of both the Glu119Val and His274Tyr mutant viruses in culture, [60] it has never been seen in clinical isolates after ferrets. [75] The Glu119Val virus grew to similar titres in both the zanamivir treatment.

donor and contact animals. Although 100-fold more virus was A second neuraminidase mutation Glu119Val, which gives needed to infect the donor animals with the His274Tyr mutant 100-to 500-fold resistance, [31, 68] has been selected after exposure virus, and virus replication was delayed by 1 day, virus grew to the to oseltamivir, in vitro in the N9 virus, and from clinical isolates in same titres in both donor and contact animals. All mutant viruses N2 viruses. [31, 67, 69] Mutant viruses are resistant only to oseltamivir. retained their genotypes after transmission. Although viruses carrying mutations at Glu119 to Gly, Ala or Asp To date, there has been no documentation of the spread of a have been generated after passage in zanamivir in vitro, [70] no resistant virus to a contact in humans, in contrast to that observed viruses with mutations at Glu119 have been isolated in patients with amantadine-resistant viruses during amantadine treatment. treated with zanamivir.

However, these recent results suggest that, despite the compro-A His274Tyr mutation has been isolated only in N1 viruses. mised nature of these viruses, there are concerns about their This mutation confers about 400-to 600-fold resistance, [68] and transmissibility in humans. has been seen in both cell culture passaged virus and in clinical isolates after exposure to oseltamivir. Resistance is specific to oseltamivir. There is a report of an oseltamivir-resistant 5. Post-Marketing Data His274Tyr neuraminidase mutant virus being isolated from an immunocompromised patient treated with amantadine, While there are many data published on the clinical trials with oseltamivir, rimantadine and zanamivir. [71] The patient continued neuraminidase inhibitors, Vogel [76] recently published an evaluato shed the resistant virus for at least 12 months. This virus was tion of the use of neuraminidase inhibitors in the normal clinical still susceptible to zanamivir.

setting. Fifty-five patients treated with zanamivir and one treated A recent report from a paediatric study in Japan also describes with oseltamivir observed a rapid drop in temperature of 1.7°C an Asn294Ser mutation in an H3N2 virus. This mutation has not within 12 hours. A 96-year-old patient demonstrated an imprespreviously been seen either in laboratory generated resistant visive improvement within 6 hours. The duration of illness was ruses or in clinical isolates and conferred about a 300-fold dedecreased by 45% and administration of antibacterials was decrease in sensitivity to oseltamivir. [31] creased by 32% compared with patients presenting with influenza Resistance in influenza B viruses has not been seen in previous-infection prior to the availability of neuraminidase inhibitors. ly healthy individuals. However, a virus with a drug-resistant Fibrinogen levels were also lower in patients treated with neuraminidase has been isolated from an oseltamivir-treated im-neuraminidase inhibitors than in patients without specific antiviral munocompromised patient infected with influenza B virus.

therapy. The author suggested that the prevention of high fibrino-TheAsp198Asn mutant neuraminidase has reduced sensitivity on-gen levels by early antiviral treatment may have an impact on the ly to oseltamivir, not zanamivir. [72] This mutation has not been incidence of myocardial infarction in patients with established seen in vitro.

coronary atherosclerosis.

In another study, military conscripts treated with zanamivir for 7. Conclusions naturally acquired influenza virus infection had a rapid reduction in virus load, up to 2.5 days of reduction in median time to Zanamivir and oseltamivir are two potent and specific inhibialleviation of symptoms, and 43% reduction in complications tors of all strains of influenza virus. Unfortunately, the focus on requiring antibacterials. [77] As well as being of benefit to the the number of days of symptom relief provided has detracted from treated patient, reduced viral shedding could result in a lower rate the importance of their impact on the severity of the symptoms. of infection spread, which would be important in a closed commu-

The greatest benefit is derived if patients are treated as early as nity. The Diskhaler ™ 1 was rated as easy or very easy to use by possible and for those presenting with severe symptoms or in high-99% of patients at the end of therapy. risk groups. Both are effective therapeutically and prophylactical-Faced with the difficulty of knowing whether the patient has ly; approval for ages and use varies in different countries. influenza and if antiviral therapy is appropriate, analysis by Smith Zanamivir has some potential advantages over oseltamivir inand Roberts [78] found that antiviral treatment of influenza without cluding acting rapidly, as it is delivered at the site of infection, rapid testing is economically reasonable in febrile patients with having no adverse effects compared with placebo, and no docutypical symptoms during the influenza season. The selection of mentation of resistance in previously healthy patients. In contrast, antiviral therapy depended on the likelihood of influenza A virus after oseltamivir administration there is a delay of 3-4 hours to infection, patient age and differences in medication cost.

reach therapeutic levels at the site of infection, some patients Zanamivir treatment was considered the most effective strategy, experience nausea and vomiting and there is a low level of minimising both quality-adjusted days lost and illness days. In the resistance observed. However, psychologically, oseltamivir has UK, the National Institute for Clinical Excellence has recommendthe greatest advantage, being orally administered. Despite surveys ed the use of neuraminidase inhibitors for elderly and at-risk showing satisfaction of patients using the Diskhaler ™ device for patients who present with influenza-like illness within 36 hours of zanamivir, [37, 38] it is far easier to prescribe a tablet, and consesymptom onset. As there was a concern that the cost of quently oseltamivir has by far the greatest share of the market. For neuraminidase inhibitors could have a significant impact on the young children and for infirm or frail elderly, oseltamivir would be national health budget, Da Silva et al. [79] sought to determine, the drug of choice. during a confirmed influenza outbreak, the proportion of eligible patients who presented in time to benefit from treatment with a With the ever-present threat of a pandemic there is a critical neuraminidase inhibitor. Only 20% of such patients, rising to 47% role for the immediate availability of neuraminidase inhibitors to in out-of-hours centres, consulted in time to benefit from treatlimit spread. However, unless there is wider acceptance in the ment. Since children play a critical role in the spread of influenza community of a role for these drugs in the treatment and prevenin the community and households, targeting children for treatment tion of influenza, there will not be sufficient stocks available to and prophylaxis with the inhibitors could play an important role in have any impact. Governments are willing to spend hundreds of further limiting the spread of influenza in the community. millions of dollars on vaccination and in advertising the vaccination campaigns, yet their reluctance to fund neuraminidase inhibi-6. Effect on Immunity tors means that they are not seriously addressing approaches to limiting spread of influenza once an outbreak has occurred. If Although vaccination is the primary method for prevention of governments were serious about trying to limit the impact of influenza, once there is an outbreak in an unvaccinated communiinfluenza in the community, they should promote greater public ty, vaccination cannot provide protection sufficiently rapidly.

awareness of the symptoms of influenza and the availability of the Neuraminidase inhibitors do not interfere with the development of drugs for both treatment and prevention. an immune response to killed influenza virus vaccines; [80, 81] therefore, they should have the potential to provide protection during the 2-to 4-week period before full immunity is induced. Since the Acknowledgments drugs do not prevent the initial virus replication, but prevent subsequent virus release, when used therapeutically, an immune No external sources of funding were used to assist in the preparation of this response develops in treated patients. Even in patients treated review. The author's institute was involved in the discovery of zanamivir. The author has previously received funding from GlaxoSmithKline for drug resisprophylactically, although asymptomatic, many develop an imtance studies on neuraminidase inhibitors. mune response, thus providing protection against reinfection. 1 The use of trade names is for product identification purposes only and does not imply endorsement.

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