By Ilene

Tamiflu (oseltamivir) has been a big seller this year, with global sales soaring to $1.93 billion in the first nine months of 2009, a 362 percent increase. Driven by the threats of a swine flu pandemic, governments have been stockpiling the drug to the tune of $1.32 billion. (Efficacy of Roche’s Flu Drug Tamiflu In Doubt)

According to David Phillips:

Allegations have surfaced that Swiss drug maker Roche has misled governments and physicians alike on the efficacy of its popular drug Tamiflu in preventing complications, such as hospitalization from pneumonia or death, in otherwise healthy people afflicted with the flu — seasonal or the H1N1 (swine flu) version. If the company is unconvincing in refuting such claims, more than its reputation could be sullied. Leveraging global concerns over avian and swine flu, Roche has seamlessly raised awareness of the purported need to treat the complications associated with the seasonal flu too. The company has successfully challenged conventional wisdom — that “rest and aspirin” be the preferred treatment option for seasonal flu — with marketing campaigns that resonate with reassuring efficacy claims for Tamiflu (oseltamivir): reductions in hospital admissions, secondary complications (including bronchitis, pneumonia, and sinusitis), and respiratory tract infections (requiring antibiotics) in otherwise healthy individuals of 61 percent, 67 percent, and 55 percent, respectively, according to Tamiflu media updates. A scarcity of published data in the medical literature motivated the nonprofit research group Cochrane Collaboration to investigate — and verify — Tamiflu’s alleged efficacy claims, particularly on the drug’s effect on the risk of hospital admission and complications in otherwise healthy people with influenza. The Cochrane review and a linked investigation undertaken jointly by the British Medical Journal and the local Channel 4 News cast doubt on the efficacy and safety of Tamiflu — and also raises disturbing questions on the drug’s promotional and marketing activities condoned by regulators on both sides of the Atlantic. Investigators disclosed that an often cited meta-analysis used as evidentiary support was based entirely on ten trials funded by Roche, only two of which were published in peer reviewed journals. The Cochrane reviewers could find no independently funded trials of Tamiflu for healthy adults. Troubling, too, former employees of the medical communcations company hired by Roche were alleged to have ghost written some of the manuscripts…. Continue reading Efficacy of Roche’s Flu Drug Tamiflu In Doubt .>>

As noted previously, in Swine Flu Virus: Changes and Consequences, the swine flu virus may be headed for greater degrees of tamiflu resistance as the genetic sequences conferring resistance (represented by the "H274Y" marker) are being passed to drug-sensitive viruses in a process called recombination. This has happened before. Tamiflu resistance was fixed in the seasonal H1N1 flu virus through this same process. Moreover, the H274Y marker is occurring along with changes in the receptor binding domain of the virus. Receptor binding domain changes appear to increase the potential severity of the infection. As Dr. Henry Niman reports in Silent Spread of Tamiflu Resistance in Texas, Recombinomics Commentary, Dec. 16, 2009:

The recent reports by WHO and the CDC on the increased detection of H274Y [i.e. genetic marker for tamiflu resistance] in pandemic H1H1 has led to concerns that Tamiflu resistance is spreading and will become fixed, as happened to H274Y in seasonal flu. These concerns were increased by the recent report of transmission of H274Y to seven healthy students during a July train ride in Vietnam… The associate of H274Y with a receptor binding domain change raised concerns, because the fixing of H274Y in seasonal H1N1 also involved a receptor binding domain change,... The latest ECDC report on influenza also noted that the H274Y sequences from the Netherlands also were mixtures and suggested H274Y was due to spontaneous mutations within the patient. However, there has been no evidence present to support spontaneous generation. There have been several reports of H274Y in samples that were from patients who initially tested as wild type, but the detection of H274Y was shortly after the start of Tamiflu treatment, indicating the H274Y was detected because of selection of a minor population already present prior to treatment…

Dr. Niman continues to assemble pieces of the genetic puzzle pointing to higher numbers of the H274Y marker being found in swine flu viruses circulating throughout the world. Today, he analyzed the sequences of viruses released in Japan and found the following:

Japan H1N1Tamiflu Resistance Sequences Signal Fixing Start

December 19, 2009

The National Institute of Infectious Diseases in Japan has released 68 full sets of H1N1 sequences from Japan. A few sequences were collected in the spring and early summer, but most are from the late summer and fall and include eight sequences with H274Y. Most of these Tamiflu resistance sequences are among the more recent isolates, signaling the recent spread of Tamilfu resistance…Additional sequences from Japan as well as other CDC sequences at GISAID indicate this sub-clade was widespread in both countries and H274Y was silently spreading. The recently released sequences from Japan and the United States identified addition sub-clades with multiple isolates with H274Y suggesting spread via minor species was common and was likely linked to the recent explosion in H274Y sequences announce by the WHO and CDC. Today the weekly CDC update (week 49) described 15 new H274Y isolates which raised double of such isolates announced in the US in the past 5 weekly reports. Similarly, 13 such isolates have been discovered recently in the Netherlands and recent H274Y cases in the US (Utah) and Netherlands have been fatal. In addition, H274Y and D225N have been detected in the same host in the US (Illinois) and France and both cases were fatal. There have also been three examples of H274Y paired with D225E, but the number of such sequences as minor species is likely to have been much higher. The recent dramatic rise in Tamiflu sequences is defining the start of the fixing of H275Y in pandemic H1N1, as happened in seasonal flu. This fixing will be facilitated by the drop in cases, which eliminates competing genomes… The drop in flu activity in the US and Japan will allow for the emergence of H274Y, which could happen in early 2010 when a new peak of activity forms. Thus, these recent changes increase causes for concern.



In case you missed my recent article on the evolution of the flu virus, I’ll reprint it here. It helps in understanding the evolution of the flu virus, which is important in assessing the future possibilities of the H1N1 pandemic.

By Ilene with guest expert Dr. Henry Niman

Background

Dr. Henry Niman heads the research company Recombinomics Inc. Recombinomics has a small group of researchers who analyze the sequence data from viral samples isolated from patients diagnosed with swine flu. Its website is a terrific place to find the newest information available.

Dr. Niman has kindly been answering my questions regarding the H1N1 virus, its evolution, and the implications regarding the spread of disease. Because the terminology may be unfamiliar, a brief introduction may be helpful towards better understanding both the H1N1 virus and the swine flu disease.

Recombination

Flu viruses, including the H1N1 varieties, are known for quickly changing genetically. Recombination is the driver of rapid molecular evolution, a process whereby small bits of genetic information pass between viruses so a virus may quickly acquire a genetic variation that has previously evolved and already exists in the viral reservoir (the pool of viruses circulating in a population). Unlike sporadic mutations, recombination reflects the acquisition of genetic material that has withstood the Darwinian test of time. Compared to sporadic mutation, recombination is a quicker, non-random mechanism for genetic change.

Changes in the H1N1 viral genome are natural. The viral reservoir consists of wild-type viruses (the predominant viruses) and low levels of variants carrying a variety of different sequences called “polymorphisms.” While recombination is not the currently favored theory regarding how flu viruses evolve, Dr. Niman believes it is the correct theory. The theory of recombination as a mechanism for genetic change has led to accurate predictions about how the flu virus would evolve as infection rates increase. As the size of the viral reservoir continues to expand, viruses with genetic differences, “polymorphisms,” become more evident.

Ukraine Outbreak

The outbreak in Ukraine was initially described in many media reports as a new lung-blackening “mystery disease,” leading to many false and misleading Internet stories. According to Dr. Niman, it was clear from the start that H1N1 was killing an unusually high number of previously healthy young adults… (See Flu News: D225G Follow-up )

Dr. Niman wrote a number of commentaries on the rising death toll and the need to make the sequences public. He predicted the deaths would be associated with a receptor binding domain change in the wild-type H1N1 virus (the predominant virus) to a variant form, characterized by the D225G genetic marker. Wild-type H1N1 has a D (the symbol for aspartic acid) at position 225 of the viral protein Hemagglutinin (HA), and is referred to as “D225.” The function of the HA protein is to bind viral particles to susceptible cells in the host animal. The variant protein, D225G, has a change in position 225, where the amino acid glycine (G) replaces aspartic acid. Hemagglutinin is one of two surface proteins projecting out from the surface of the virus.

The D225G marker represents a change in a single nucleotide in the virus’s genetic sequence encoding the HA protein. The amino acid change in the virus’s receptor binding protein allows the virus to bind receptors in lung tissue with greater affinity than the more usual binding in the upper airways, where the wild-type D225 protein binds. Theoretically, this may confer greater virulence, potentially leading to more severe disease as the virus invades deeper in the lungs. This change was also seen in the 1918 flu pandemic, in some, but not all, cases.

Dr. Niman reported that the genetic marker D225G was identified in lung tissue of patients who died from the effects of cytokine storms. A cytokine storm , or hypercytokinemia, “is a potentially fatal immune reaction consisting of a positive feedback loop between cytokines and immune cells, with highly elevated levels of various cytokines. The primary symptoms of a cytokine storm are high fever, swelling and redness, extreme fatigue and nausea. In some cases the immune reaction may be fatal.” Wikipedia . “ Cytokines are any of a number of substances that are secreted by specific cells of the immune system which carry signals locally between cells, and thus have an effect on other cells.” Wikipedia. The higher than usual death rate in the 1918 flu pandemic appears to be a consequence of the virus’s ability to provoke these cytokine storm reactions in patients.

Dr. Niman’s Unifying Theory

According to Dr. Niman, the most likely explanation for the concurrent emergence of the D225G and Tamiflu (or oseltamivir)-resistant variants in multiple regions is that the swine flu viruses circulating are not a homogeneous strain, but a mixture of a wild-type strain with a variety of less common variants. Less common variants include viruses with the D225G genetic marker and also viruses containing genetic sequences (H274Y) conferring Tamiflu resistance. The H274Y marker represents the genetic change in the neuraminidase gene (NA gene) which encodes the NA protein. The H247Y change, from the amino acid histidine to tyrosine in the NA protein, leads to Tamiflu resistance.

There are enough D225G variants and H274Y variants in the viral reservoir to act as “donor sequences” so the genetic changes represented by these markers can jump from one virus to another, leading to their simultaneous detection in many locations. Detection increases as the viral reservoir expands, along with D225G and H274Y variants (more viruses, greater numbers of people infected), so greater numbers of the non-wild-type viruses show up in flu cases. Dr. Niman believes the D225G marker is not adequately represented in the flu database because this variant is not easily detected in nasopharyngeal swabs, due to its preferential binding in lungs.

The background presence of the D225G and H247Y variations in the H1N1 virus pool explains why cases with the D225G and H247Y markers are found throughout the world, in mild as well as severe cases. The presence of viruses with these marker should not be seen as an all-or-nothing phenomenon. Rather, smaller number of these viruses appear to co-exist with the more prevalent wild-type viruses in the viral reservoir.

Theoretically, if a D225G subclone takes hold in the lungs and expands, it can cause a more severe flu. While the D225G marker may increase the virus’s virulence, the receptor binding profile is only one of a number of factors influencing the severity and outcome of infection. Other factors include the viral load (how much virus a patient is exposed to), the patient’s immune system (does the patient have antibodies to the virus?), and other characteristics of the infecting viruses–e.g., the presence of genetic marker H247Y, resulting in tamiflu resistance. In addition, the severity of an outbreak will be influenced by the virus’s transmissibility, the more transmissible, the greater numbers of people infected. Once infected, however, our immune response can make the difference between a mild or lethal disease course.

We could have seen this coming, though. This type of virus emerged in the US in 1998 and has since become endemic on hog farms across North America. Equipped with a suite of pig, bird and human genes, it was also evolving rapidly… By 1999, these viruses comprised the dominant flu strain in North American pigs and, unlike the swine virus they replaced, they were actively evolving. There are many versions with different pig or human surface proteins, including one, like the Mexican flu spreading now, with H1 and N1 from the original swine virus… Our immune response to flu, which makes the difference between mild and potentially lethal disease, is mainly due to the H surface protein. The Mexican virus carries the swine version, so the antibodies we carry to human H1N1 viruses will not recognise it…. See: Swine flu: The predictable pandemic?

In Dr. Niman’s opinion, a new wave of flu infections will come in early 2010, and Tamiflu resistance and receptor binding domain changes will be common. This new wave may be beginning in Ukraine currently .

See also: Swine Flu News: What is the significance of D225G? and Flu News: D225G Follow-up .

The Emergence of Tamiflu Resistance

The swine flu virus has been increasingly showing changes leading to greater incidences of Tamiflu resistance, but this being downplayed by the CDC and WHO. The CDC recently issued this CDC report :

A total of 29 cases of oseltamivir resistant 2009 influenza A (H1N1) viruses have been identified in the United States since April 2009. In specimens collected since September 1, 2009, 19 cases have been identified in the United States, including three newly identified cases since last week. The proportion of oseltamivir-resistant 2009 H1N1 viruses does not represent the prevalence of oseltamivir-resistant 2009 H1N1 in the U.S. Most cases were tested because drug resistance was suspected. All tested viruses retain their sensitivity to the neuraminidase inhibitor zanamivir. Of the 29 total cases identified, 19 patients had documented exposure to oseltamivir through either treatment or chemoprophylaxis, eight patients are under investigation to determine exposure to oseltamivir, and two patients had no documented oseltamivir exposure. Occasional development of oseltamivir resistance during treatment or prophylaxis is not unexpected. Enhanced surveillance and increased availability of testing performed at CDC are expected to detect additional cases of oseltamivir resistant 2009 influenza A (H1N1) viruses, and such cases will be investigated to assess the spread of resistant strains in the community.

According to Dr. Niman, the CDC and WHO typically use these types of announcements for propaganda. The information is factually correct, but extremely misleading. Most patients give a sample prior to treatment and in most cases CDC or WHO can determine if the H274Y marker was present prior to treatment. However, they don’t even have a category for those who are initially H274Y positive and then get treated, like the Vietnam cluster .

When there is a small population of drug-resistant viruses co-existing with the wild-type, drug-sensitive viruses, exposure to the drug kills off the sensitive population while allowing the resistant viruses to flourish. In cases testing positive for the H274Y marker, the vast majority of patients has been infected with H1N1 virus with the H274Y marker before treatment, or develops H274Y within a few days. This suggests a pre-existing sub-population is being selected. Data withheld by the CDC and WHO would likely reveal this, destroying the “spontaneous generation” theory, which the CDC, WHO and Roche promote. These propaganda pieces are not only designed to hide the true level of H274Y in the absence of Tamiflu treatment, but also hide the fact that “spontaneous” H274Y mutation is a pure fantasy. The emergence of Tamiflu resistance is NOT due to random mutation.

For a more detailed account, read Dr. Niman’s article Tamiflu Resistance Spike in US Raises Transparency Concerns, Recombinomics, December 14, 2009:

[The CDC report (week 48, above)] announces three more cases of Tamiflu resistance in the US. This number matches the increases for each of the past 3 weeks and brings the total for the past 4 weeks to 16, which is much higher than previous weeks, which usually had 0 or 1 new cases. This recent spike in cases has also been reported by WHO and raises concerns that H274Y is efficiently transmitting. Moreover, recent deaths of patients with H274Y in the US (four of ten) and the Netherlands (four of eleven) have raised concerns that patients with H274Y also have D225G, which has been associated with fatal cases in the US, Ukraine, Norway, Brazil, and France. Moreover, patients with D225G coupled with H274Y have been reported in France and the United States. However, the CDC report does address those concerns because critical data has been withheld. In week 48 the number of samples tested for H274Y spiked higher, but there is no indication of these are recent samples or an update of surveillance done on samples collected in the spring or summer. Similarly, the location of these cases or outcomes are not given, and there is no indication that patients who developed resistance during treatment are distinguished from patients who were resistant prior to treatment, but fell into the “suspect” category because they failed to respond to Tamiflu. The Vietnam cluster described in this week’s New England Journal of Medicine that would represent such cases. Seven passengers on a train developed H1N1 infections that had H274Y. However, like most patients worldwide, the resistance was not discovered until long after treatment and discharge. The infections were in July, but the first lab confirmations were in September. Although Tamiflu treatment is not effective against H1N1 with H274Y, all patients recovered, but clearly represented infections of a fit and readily transmissible H1N1 with h274Y. However, using the CDC classification system, these patients would have “documented exposure to Tamiflu” but would not be examples of resistance that developed due to treatment. Although this is the CDC’s largest category, they have yet to show a single example of a case that developed resistance after prolong treatment with Tamiflu… The failure of the CDC to report any patients who developed H274Y after prolonged treatment, and the efficient transmission of H274Y in Vietnam in July, raise concerns that the weekly reports by the CDC are carefully designed to withhold key information such as the H274Y status prior to treatment, the dates and locations of samples, as well as outcomes of patients who are H274Y positive. This lack of transparency continues to be hazardous to the world’s health.

******

Dr. Henry Niman

Dr. Niman earned a PhD in biochemistry at the University of Southern California in 1978, and as a coincidence, he and I worked in the same pathology/biochemistry laboratory at USC, separated only by time. His dissertation focused on feline retroviral expression in tumors. Working on his post-doctorate at the Scripps Clinic and Research Foundation, and later accepting a staff position, Dr. Niman began looking at making monoclonal antibodies using synthetic peptides. Data generated in 1982 demonstrated that the two technologies–monoclonal antibody and synthetic peptide technologies–could be combined. His work led to the popularity of the flu monoclonal antibody, which is widely used throughout the pharmaceutical, biotech, and research industries. He also produced a broad panel of monoclonal antibodies against synthetic peptides of oncogenes and growth factors.

Dr. Niman subsequently had a joint appointment as Instructor in Surgery at Harvard/Massachusetts General Hospital and as Research Associate at the Shriner’s Burn Center across the street from Mass General. (These were research positions – he did not teach or do surgery.) The technology developed by Dr. Niman was used to form ProgenX, a cancer diagnostic company that became Ligand Pharmaceuticals. More recently, he has been studying infectious diseases and viral evolution.

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