Rintatolimod, aka Ampligen, on Trial

By Suzanne D. Vernon, PhD
Scientific Director

Clinical trials occur in phases. A Phase I trial signifies the first time a drug or treatment is tested in humans to assess its safety, dosing and side effects. Phase I trials can be small and may involve healthy volunteers. Phase II trials generally involve 100-300 people in testing, and protocols are designed to further evaluate efficacy and safety. Phase III trials are conducted on larger groups (1,000-3,000) to confirm effectiveness, monitor adverse events and compare outcomes to commonly used treatments. Phase IV trials include studies to gather additional information on the drug’s risks, benefits and optimal use.

In 1994, promising results of a Phase II trial of Ampligen® in 92 chronic fatigue syndrome (CFS) patients were published in Clinical Infectious Diseases (1), setting the stage for a larger Phase III trial. Ampligen is a double-stranded RNA molecule. Double-stranded RNA is also found in certain types of viral genomes or as part of the viral life cycle.  Therefore, double-stranded RNA is a potent activator of the innate immune response. Ampligen was designed to mimic the immune response and defend against viral infection. These early promising results as well as reports made by manufacturer Hemispherx Biopharma at meetings and conferences and by patients taking the drug over the intervening 18 years have created a great deal of anticipation and intrigue about this particular therapy.

At long last, the results of a multi-center Phase III Ampligen trial were described in a March 14, 2012 publication in PLoS ONE (2).It is important to note that this Phase III trial actually took place between 1998 and 2004. It was a double-blind, placebo-controlled, randomized clinical trial that enrolled 234 CFS patients at 12 different clinical sites. Patients were intravenously infused twice weekly for 40 weeks with either Ampligen or a placebo solution; this was Stage 1. After the 40-week blinded period, those who received placebo were switched to Ampligen infusions for 24 weeks. The individuals who received Ampligen for 40 weeks continued with the drug for 24 more weeks. This period of treatment was referred to as Stage 2.

A number of measures were collected before the study, during and after it to judge the effectiveness of the drug. The primary measure of success (known as a primary outcome) was the duration of exercise tolerance testing an individual could perform. The exercise tolerance test chosen by the drug manufacturer for this study involved a treadmill test. It had a number of stages in three-minute increments, during which the speed and the incline of the treadmill were increased. For this kind of test, every incremental second requires more effort than the last. In addition to the time on the treadmill, subjects rated their perceived exertion. The secondary outcome measures were 1) the number of medications taken for CFS symptoms during the 40-week trial and 2) measures of function including the Karnofsky Performance Scale (KPS), activities of daily living (ADL) score, and the vitality and general health perception sub-scores from a standardized tool called the Short-Form 36 (SF-36).

The CFS patients included in this study were described as being severely ill, had been ill a minimum of one year and met Holmes (1988) and Fukuda (1994) criteria for CFS. (This study predates the 2003 Canadian criteria for ME/CFS.) They had to have a KPS score between 40 (disabled: unable to care for self) and 60 (requires occasional assistance but is able to care for most needs). There were numerous other inclusion and exclusion criteria for entry into the trial, outlined in Table S1 of the Supporting Information.

Prior to being randomized to drug or placebo, the patients were divided into two groups: those who could complete nine minutes or less on the treadmill and those who could complete more than nine minutes but fewer than 18 minutes of exercise on the treadmill. The group receiving drug and placebo were similar in age (43 years), weight (167 pounds), age at CFS onset (34 years), duration of CFS symptoms (10 years), time from CFS diagnosis (six years) and ethnicity (91-93 percent Caucasian). There were fewer women receiving drug (67.5 percent) than placebo (77.8 percent), but this difference was not statistically significant.

Adverse events — side effects experienced by those in the study — were roughly the same in both groups, with 99 percent of patients in the Ampligen group reporting an adverse event and 97 percent of placebo-treated individuals also reporting adverse events. There were three serious adverse events in each study arm judged to be unrelated to the study drug. There were two deaths in Stage 2 related to non-exclusionary pre-existing conditions. According to the PLoS ONE report, “…neither [death] in the opinion of the principal investigators at the clinical sites was related to the study drug.”

When the data were analyzed, it was shown that receiving Ampligen had a positive and significant effect on increasing exercise tolerance. The group that received Ampligen for 40 weeks increased from 583 seconds at baseline to 691 seconds. The group that received placebo for 40 weeks also increased, but not as much; they went from an average of 587 seconds at baseline to 614 seconds at 40 weeks. When this group received Ampligen for 24 weeks during Stage 2, they experienced a further increase equal to the first group. The group that continued with Ampligen sustained the increased tolerance of exercise.

Therefore, the increase in time on the treadmill observed with Ampligen – both in the group receiving the drug for the entire 40-week period and in the placebo group that began receiving the drug at week 40 – was sufficient to demonstrate the efficacy of Ampligen to achieve the primary outcome measure. The improvement in exercise tolerance seen with Ampligen is similar to or slightly better than other drugs used to treat chronic conditions with exertional fatigue (e.g., chronic congestive heart failure, pulmonary arterial hypertension, etc.).

The CFS patients who received Ampligen reported using fewer symptom-relieving medications compared to those who received placebo. Sixty-eight percent of the Ampligen-treated group reduced their use of other medicines, while 55 percent of the placebo-treated group did. (The medications used during the study are listed in the Supporting Information.) The other secondary performance endpoints are also reported as significantly different in those receiving drug compared to placebo (Table 4, above). However, it is not clear that these statistical differences actually indicate whether the patients felt and functioned better.

The data described in this Phase III clinical trial of Ampligen are important because the primary endpoint measure of exercise tolerance was achieved. While a minute or so of increased exercise tolerance may not seem like much for a healthy person, it is likely a significant accomplishment for a person with CFS, especially using this kind of testing protocol. It is also important to know if this increase in exercise tolerance time translated to patients feeling better; that is a measure that FDA will consider if and when Hemispherx submits a revised new drug application (NDA) for Ampligen. There might be data to support improved patient function, but the PLoS ONE report doesn’t lend itself to this kind of interpretation. A Karnofsky Performance Scale change of five points – from an average of 50 to an average of 55 – is still not an acceptable level of function and it doesn’t rise above the level required for entry into the study. A vitality score change from 5 to 10 on a 100-point scale still results in a very low level. The placebo group’s vitality score was 10 at the beginning of the study and 10 at the end of 40 weeks. According to the publication, the Ampligen-treated group participants’ general health perception didn’t change at all; it was 20 at the beginning of the study and 20 at the end, whereas the placebo group’s score went from 20 to 25 at week 40.

How could this be? Over the more than 20 years that Ampligen has been used by a handful of expert CFS clinicians to treat CFS patients, there have certainly been anecdotal treatment successes. Certain CFS patients appear to respond fabulously to Ampligen and go from being bed- or wheelchair-bound to regaining near-normal physical function while they are being treated. However, there are also anecdotal reports of Ampligen treatment failures. What do we know about the CFS patients that respond — or not — to Ampligen?

The PLoS ONE paper describes a conventional clinical trial design and traditional statistical assessment. We are now in the post-genomic era with sophisticated tools that can be used to aid clinical trial design, patient selection, understanding mechanism of action of a compound and systems pharmacology. This study was initiated at a time when relatively little was known about CFS and before there was an ability to use powerful genome information for targeted clinical trials. It used crude measures like the Karnofsky Performance Score that in hindsight aren’t likely to discriminate improvement very well.

There are hints of promise for Ampligen — especially in the anecdotal treatment success stories – but those are difficult to uncover in data where all participants are lumped together. Hemispherx Biopharma could learn from the experience of the lupus drug Benlysta, approved by the FDA in 2011. Benlysta’s drug developer delved more deeply into the “lumped” data to identify specific subgroups of lupus patients who responded to that drug. This subgroup analysis led to FDA approval. The PLoS ONE paper recognizes the possibility that gene expression profiles may help define subgroups:

“The availability of diagnostic assays, based on gene expression profiles, may provide a prognostic genetic signature that will predict response, as well as the opportunity to develop synergistic combinations of rintatolimod and other agents to expand the therapeutic effect in this debilitating disease.”

We know CFS is heterogeneous. We know there are those individuals with CFS who benefit from Ampligen. After more than 20 years of CFS Ampligen clinical trials, there is bound to be rich data that could be used to determine the CFS “type” that responds to Ampligen. As patent holder and manufacturer of Ampligen, Hemispherx Biopharma has an ethical responsibility to patients and its investors to dig as deep as possible into the mountains of data it has gathered. Hopefully, Hemispherx Biopharma will team with experts using 21st century bioinformatics tools to conduct more sophisticated subgroup analyses as part of its next submission to the FDA. The last ruling from FDA in 2009, recommending a six-month study of at least 300 patients, suggests that it needs more convincing data than what has been presented so far.


1. Strayer DR, Carter WA, Brodsky I, Cheney P, Peterson D, Salvato P, Thompson C, Loveless M, Shapiro DE, Elsasser W, et al. A controlled clinical trial with a specifically configured RNA drug, poly(I).poly(C12U), in chronic fatigue syndrome. Clin Infect Dis. 1994 Jan;18 Suppl 1:S88-95. Abstract: http://www.ncbi.nlm.nih.gov/pubmed/8148460

2. Strayer DR , Carter WA , Stouch BC , Stevens SR , Bateman L , et al. (2012) A double-blind, placebo-controlled, randomized, clinical trial of the TLR-3 agonist rintatolimod in severe cases of chronic fatigue syndrome. PLoS ONE 7(3): e31334. doi:10.1371/journal.pone.0031334 URL: http://www.plosone.org/article/info:doi%2F10.1371%2Fjournal.pone.0031334 (Open Access)

Media coverage of this study:
Press release issued by Hemispherx Biopharma  (Mar. 19, 2012)
“Is Hemispherx BioPharma, Inc. Finally Seeing the Light at the End of the Tunnel?” SmallCapNetwork.com (Mar. 19, 2012)

Suzanne D. Vernon, PhD, is the Solve ME/CFS Initiative’s scientific director. She has nearly two decades of experience as a microbiologist.

March 21, 2012