This summer’s Olympics are set to be the toughest yet for substance-abusing athletes. Traditional tests are already identifying cases like Morocco’s Mariem Alaoui Selsouli, a gold-medal hopeful in the 1,500 meters who is now barred from competing in London after testing positive for the banned diuretic furosemide, which can help mask the use of steroids and other illicit substances. And this year scientists are adding more powerful techniques to their arsenal to catch those athletes that have begun to tailor their doping strategies to avoid detection.
The new tests cast a wider net by “using lots of different strategies,” explained Phil Teale, chief scientist in Medication & Doping Control at the drug-testing laboratory HFL Sport Science in the United Kingdom, “moving away from focused approaches” that can only detect a pre-defined set of substances.
Researchers have historically used mass spectrometry to identify anabolic steroids and other prohibited compounds, but this technique requires a preexisting knowledge of the drugs’ structures. This allowed “designer” steroids, like tetrahydrogestrinone (THG), which have the same effects as other steroids—like increasing speed or boosting muscle mass—but a slightly different structure, to evade detection, Teale noted. But the London Olympics lab, a joint effort between pharmaceutical company GlaxoSmithKline and the Drug Control Centre at King’s College, London, has several mass spectrometers capable of recording all sample data, Teale said, which will allow officials to retroactively screen samples for newly-identified performance enhancing substances.
Researchers are also investigating cell-based assays that can give information about steroid abuse even if scientists don’t know which steroids to test for. One such method relies on a yeast-based reporter system, in which ß-galactosidase is turned on when anabolic steroids activate the androgen receptor. In a recent study, urine samples containing a range of anabolic steroids—including THG—prompted expression of ß-galactosidase, while non-androgenic steroids like estrogen did not. Another study showed that the yeast reporter system could detect a single steroid dose 2 weeks after ingestion, while classic mass spectrometry tests only detected the drug for the first 6 days.
Other strategies for doping, like injecting human growth hormone (hGH), which some athletes believe will boost performance by building muscle, can also be difficult to detect directly. The test for hGH used in the 2008 Olympic Games measures changing ratios of growth hormone isoforms to detect injection of recombinant hGH, which mimics only one particular natural isoform. But it can only detect recombinant hGH injected with the last 24 hours, said Peter Sönksen, emeritus professor at King’s College, London. Such a test is thus ineffective at detecting long-term hGH abuse, and hGH must be taken over months to show effects. But a new biomarker assay, approved just a few weeks ago by the World Anti-Doping Agency (WADA), is making its Olympic debut at the London Games. It monitors unexpected increases in insulin-like growth factor-1 (IGF-1) and collagen proteins that are produced in response to hGH to reveal abuse of human growth hormone (hGH), and can detect artificial hGH use over 2 weeks after injection, said Sönksen, who helped spear-head development of hGH biomarker testing in the 1990s.
Another common performance-enhancing drug is erythropoietin (EPO), which boosts the production of red blood cells, allowing athletes to increase the oxygen-carrying capability of their blood. “If you go to [high] altitude, you turn up EPO, but you also turn up other genes,” explained James Rupert, who studies EPO at the University of British Columbia. But if only EPO levels are elevated without the accompanying suite of altitude-induced genes, it suggests artificial administration of EPO, perhaps through injection of the recombinant protein, he said. Although this research has yet to result in an anti-doping test, “the principle is sound,” said Rupert, who hopes to see an EPO test included among Olympic anti-doping tests in the future. Another new EPO-detecting strategy being researched by Swedish scientists relies on different profiles of carbohydrate modifications on recombinant versus endogenous EPO. If successful, this test will be able to detect smaller amounts of recombinant EPO than current methods.
Scientists are also searching for indirect evidence of doping. One popular method to boost red blood cells is by transfusing an athlete with their own blood, previously removed and stored. Autologous blood doping, which has the same benefits as EPO, “is impossible to detect,” explained Carsten Lundby, a cardiac physiologist at the University of Zurich. It doesn’t alter an athlete’s hormone profile or DNA sequences (like blood from a donor might)—changes that would be measurable by traditional tests. But scientists are trying anyway. One new technique, for example, doesn’t even look at blood; it looks at urine. Chemical compounds used in the plastic bags that store blood, known as phthalates, can leach into the samples, and transfusing the blood back into an athlete will introduce the unusual compounds into the body. Researchers in Spain and Hungary are devising a technique to measure metabolic byproducts of phthalates in urine as evidence of blood transfusion.
An added difficulty in determining whether measurements of potential doping agents, like hormones, are truly elevated in an athlete’s blood is that levels can vary widely between individuals, and elite athletes may not have such “normal” levels to begin with, said Rupert. Instead, it’s becoming more common to take multiple samples of an athlete’s blood and urine over time, building a “biological passport” that gives doping officials a standard normal range for each person. Only variations beyond an individual athlete’s normal range would raise a suspicion.
Researchers are “investigating a really broad range of methodologies,” said Teale. With the current tests and new strategies under development, he explained, “we really have a chance to interrogate [athletes’ samples] and dig in.”