Adenoviruses are the most common carrier systems used in gene therapy studies to transport therapeutic genes into cells in the body. However, in the USA two years ago, Jesse Gelsinger, an 18-year-old patient, died suddenly of organ failure after direct injection into his bloodstream of attenuated adenoviruses used as a “gene taxi”. The exact molecular cause of his death remains unexplained. Now, a report of a hitherto unobserved immune reaction to adenoviruses has now been published by a research team headed by Dr Günter Cichon from the Humboldt University of Berlin, at the Max Delbrück Center for Molecular Medicine (MDC), Berlin Buch, and Prof. Reinhard Burger from the Robert Koch Institute, Berlin, in the January issue of the journal “Gene Therapy” ( Vol. 8, Issue 23, 2001, pp. 1794 – 1800)*. In the laboratory, large quantities of adenoviruses as used in gene therapy triggered an unexpectedly strong activation of the so-called complement system. The complement system consists of a group of proteins which circulate in the blood and act as an initial protection system against the threat posed by infectious pathogens. In order to increase patient safety, the authors suggest that in the future the status of the complement reaction be measured by a simple test before gene therapy is administered.

The task of the complement system proteins is to alert the body’s immune defenses. An unnaturally occurring, sudden massive trigger – for example, the sudden appearance of a high concentration of virus in the blood  – can trigger a dangerous complement revolt. “This immune overreaction can produce severe damage to organs and threaten patients’ lives”, explained Prof. Burger, senior author and an immunologist at the Robert Koch Institute in Berlin. Such fatal attacks by the complement system are known to occur in trauma patients after serious accidents, burns or after transfusion of blood of the wrong blood group. When this happens, signs of a massive inflammatory reaction appear in the vessel walls of the liver, lungs and kidneys, which can lead to multiorgan failure in some cases.

“So far, the role of the complement system in the side-effects that happened to Jesse Gelsinger and encountered in other adenoviral gene therapy studies has not been discussed in the scientific literature”, explained the physician, Dr.  Cichon, from the Humboldt University at the MDC, Berlin. In their laboratory investigations, the authors mixed raising concentrations of a number of adenoviruses - including serotype Ad5 frequently used in gene therapy studies  - with the blood plasma of 18 volunteer donors. Then, they analyzed the concentrations of an indicator of complement activation, protein C3a-desArg, one of the so-called anaphylatoxins. To their surprise, the researchers found that virus concentrations that are also achieved in the blood of gene therapy patients during gene therapy, caused massive activation of the complement system. This occurred in all those subjects who had previously had contact with natural adenoviruses at some time in their lives and exhibited evidence of antibodies to the pathogens.

“We must assume that there is significant activation of the complement system as a consequence of infusing high doses of adenoviruses into the human bloodstream”, suggested Dr. Cichon as an explanation of their findings. As far as the fatal gene therapy treatment was concerned, Jesse Gelsinger received an infusion of 300 thousand million (3x10¹¹) virus particles per kilogram body weight via the hepatic artery. This is equivalent to a viral dose of 7.5 thousand million (7,5x10⁹) particles per milliliter blood plasma. In the case of an equivalent viral dose in isolated plasma in a test-tube, the researchers found an astonishing 3000 nanograms C3a-desArg protein per milliliter blood plasma following the reaction of the complement system. In normal circumstances, the value is less than 150 nanograms per milliliter. From clinical studies involving severely injured patients (particularly after burns and accidents involving massive tissue destruction e.g. following serious road traffic accidents), it is known that intense complement activation can occur. If the concentration in blood reaches a peak of 1000 ng C3a-desArg/ml plasma, there is a markedly increased risk of the development of uncontrolled inflammatory reactions which can lead to liver and kidney failure as well as seriously affecting lung perfusion. The induced complement activation following administration of adenovirus under laboratory conditions is 3000 ng C3a-desArg /ml plasma, about three-fold higher than the clinically measured peak value.

The researchers stress that at the moment it is not clear how the data obtained in vitro can be directly extrapolated to gene therapy patients. However, because adenoviruses can act as promising vectors for the gene therapy of cancer, great attention will need to be paid to the complement system as a potential trigger of side effects in future clinical studies. “The Jesse Gelsinger case shows that it is very dangerous to administer high doses of virus”, says Dr. Cichon. Prof. Burger added his own comment: “Our new measuring system acts like a sort  of probe. It signals if there is a chance of patients suffering life-threatening side effects of gene therapy with adenoviruses.” It may be helpful to give drugs that are  known to be able to prevent overactivation of the complement system.

However, the researchers believe that it is important to win patients’ confidence as far as molecular therapy methods are concerned. The discussion of potential side effects and the development of methods to avoid them will form an important basis for this. At the moment, the side effects of gene therapy studies are rarely discussed at a professional level since the detailed  clinical data that could provide the basis for such discussion are rarely published because of the involvement of commercial companies. The American health authorities are presently trying to alter precedents in order to resolve this dilemma, an attempt, which could act as an example to European health authorities.

* Complement activation by recombinant adenoviruses

Barbara Bachtler

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