Triple whammy may inhibit Tamiflu as bird flu weapon

The failure of the anti-influenza drug Tamiflu to save the lives of patients with established H5N1 avian influenza infections may be a consequence of the drug's design, mode of delivery, and the uniqueness of the virus itself, according to experts.

Many governments around the world have spent tens of millions of dollars stockpiling supplies of the US-developed neuraminidase inhibitor, in preference to the original, Australian designed anti-influenza drug Relenza.

Last month Vietnamese doctors described Tamiflu (oseltamavir phosphate) as "useless" as a treatment for an established H5N1 infection, and the World Health Organisation said the drug had "not been widely successful in human patients". Three siblings from a Turkish family last week became the first European fatalities attributed to avian influenza.

Former CSIRO x-ray crystallographer Dr Peter Colman, whose research on the structure of the virus' neuraminidase enzyme led to the development of Relenza, said Tamiflu differs subtly from Relenza in the way it binds and inactivates the virus' neuraminidase coat protein.

The two drugs also differ fundamentally in their mode of delivery. Relenza is inhaled directly into the respiratory tract, via GlaxoSmithKline's patented DiskHaler oral inhaler -- the drug is not orally available. Tamiflu is orally administered, so must transit the digestive tract, bloodstream and the liver before reaching its site of action in the respiratory tract and achieving a protective concentration. Consequently, it must be used at a much higher dosage.

Contrast

Dr Anne Moscone, an expert on neuraminidase inhibitors at Cornell University, contrasted the activity of the two drugs in a review in the New England Journal of Medicine last September. The replication of the influenza virus in the respiratory tract reaches a peak between 24 and 72 hours after the onset of illness, she said, so the drugs need to be administered as early as possible.

During the deadly Spanish flu epidemic of 1918-19, which killed an estimated 40 million people on six continents, healthy young adults died within six hours of exhibiting the first symptoms of infection. Recent research suggests this age cohort was particularly vulnerable because their healthy immune systems responded by flooding their bodies with cytokines -- cell signalling compounds -- resulting in asphyxia due to massive leakage of fluid from distressed cells into the lungs.

Moscone noted that 10 to 20 per cent of Relenza's active ingredient, zanamavir, reaches the lungs, and becomes highly concentrated in the respiratory tract. "The concentration of the drug in the respiratory tract has been estimated to be more than 1000 times as the 50 per cent inhibitory concentration... for neuraminidase; in addition the inhibitory effect starts within 10 seconds -- two favourable features in terms of reducing the likelihood of emergence of drug resistant variant viruses," she wrote.

Australian-born virologist and influenza expert Dr Robert Webster, of St Jude's Children's Hospital in Memphis, Tennessee, last week told Reuters that research was still required into the optimal dose for Tamiflu, and the correct dosing regime. Webster was commenting on a report that eight Vietnamese patients have died of 'bird flu' despite treatment with Tamiflu.

On Moscone's assessment, Relenza rapidly achieves 1000 times the protective concentration required at the lung surface. According to the Moscone paper, oseltamavir is readily absorbed via the gastrointestinal tract in powdered or liquid form, and esterase enzymes in the liver then convert it to its active form. It becomes widely distributed in the body, which means it can act in tissues or organs other than the respiratory tract.

For normal influenza infections, Tamiflu reduces the duration of infection by more than 30 per cent (3 to 4.3 days) and the severity of illness by about 40 per cent -- but only when administered early. "Early initiation of treatment appears to be the most important determinant of treatment efficacy," Moscone wrote.

Japanese deaths

Relenza has been less extensively tested, but initial tests on children in 1997 indicated it reduced the duration of infection by two days. Relenza appears to be well tolerated by children; the relative safety of the two drugs became an issue when Japanese researchers reported the deaths of five children treated with Tamiflu for non-H5N1 influenza.

Because Relenza is still not widely used as a treatment for established infections, it is not clear whether it would have been any more effective than Tamiflu in saving the lives of the Vietnamese and Turkish patients, or other individuals who have died of avian influenza in China, Indonesia and Thailand.

But according to Peter Colman, there are reasons to suspect that Relenza might be more efficacious than Tamiflu in treating an established infection.

Both drugs work by binding to a small pocket-like structure in the virus' neuraminidase coat protein, inactivating the enzyme so that newly assembled virions cannot free themselves from the plasma membrane of the infected cells to infect adjacent cells.

"Tamiflu requires a small change in the conformation of the (neuraminidase) protein to bind effectively," Colman said. "We know that in some strains of the virus, the drug has to do more 'work' to mould the structure of the neuraminidase molecule to itself, which manifests as a weaker binding reaction."

According to Moscone, the Roche drug requires an amino acid residue, E276, which lies within the active 'pocket' of neuraminidase to rotate before it the drug can bind to it. She said this mechanism has clinical implications: in all influenza strains that have evolved resistance to Tamiflu, the protective mutations involve this residue, or adjacent residues, that prevent the residue rotating into position for the binding reaction. Five Japanese children who died of H5N1 influenza last year despite treatment with Tamiflu were found to be infected by viruses with this type of mutation.

Relenza does not require this rotation to bind effectively, reducing the risk of resistant strains of the virus surviving Relenza therapy.

At an FDA hearing in the US last month, Roche denied any causal relationship between Tamiflu therapy and the deaths of 12 Japanese children. Japanese health authorities reported that 32 other children developed psychiatric and emotional symptoms after being treated with Tamiflu -- but it is not yet clear whether their symptoms were due to the virus or the drug.

Japan is the world's biggest consumer and stockpiler of Tamiflu; more than 35 million doses have been administered in Japan to prevent or treat influenza.

According to Colman, while Tamiflu must 'work' harder to bind to N1 neuraminidase molecules in type B influenza viruses than in type A viruses (H5N1 is a type A virus) he knows of no evidence that the binding 'penalty' was higher for the particular variant of the N1 enzyme carried by H5N1 virus strain.

"The test-tube-evidence is that both drugs inhibit all N1 type influenza viruses, and I don't know if there is anything subtly peculiar about the structure of the H5N1 neuraminidase," he said. "As to claims that doctors have been unable to get infected patients to respond to Tamiflu, there will always be an issue with how far the infection has advanced -- any drug for influenza will have to be taken fairly early to be effective."

Colman said the considerably longer time required for Tamiflu to reach a therapeutic concentration the respiratory tract, via the oral route, could be significant.

It's personal

Dr James Campbell, vice-president of operations at ChemGenex Pharmaceuticals, which specialises in developing 'personalised' cancer drugs, said it was quite likely that individual differences in the way infected patients' livers metabolise the drug, en route to the respiratory tract, could result in problems achieving a therapeutic concentration in the lungs.

ChemGenex researchers have found that a single nucleotide polymorphism (SNP) in a liver-enzyme gene results in major differences in the rate at which individuals metabolise the company's development cancer drug Quinamed.

Campbell said the fact that Tamiflu was orally administered meant it may be exposed to breakdown by the cytochrome P450 family of liver enzymes, or other liver enzymes that detoxify and clear natural poisons and drugs from the body.

Depending on which variants of the cytochrome P450 genes patients inherit, their responses to a standard Tamiflu dose, and the half-life of the drug in the body, might differ significantly.