Comparative genome analysis has changed theories about evolution and disease. Andrew West examines how it may also change ideas about drug development.

Comparative genome analysis has changed theories about evolution and disease. Andrew West examines how it may also change ideas about drug development.

The genomes of rodents, primates and humans have recently been compared for the first time, and the results may have some far-reaching consequences for the world of drug discovery.

The search for new drugs involves a labyrinth of trials where 99 per cent of the paths are a dead end. Despite millions spent on research and the involvement of some of the world’s top scientists, the successful commercialisation of drugs remains a complicated affair marred by low compound discovery and high failure rates.

Now, as well as answering some fundamental questions about mankind, genome sequencing projects are being used in conjunction with current technologies to help revolutionise the way potential cures for human diseases are evaluated. Genome sequencing involves determining the position of every chemical base in the DNA of the species being examined. This data can be used to establish the positions within the DNA strands of genes that have a specific function in the organism. By comparing animal genomes to the human genome, important genetic features, such as genes responsible for disease and immunity, can be identified and contrasted. Then, rather than discovering new drugs by trial and error, drugs can be specifically designed to target a particular defective gene. Treatments can be tailor-made for individuals and adverse reactions to drugs will be easier to predict and avoid, saving time, the high cost of unsuccessful clinical trials and the lives of thousands of laboratory animals.

But rather than validating current testing techniques as hoped, the publication of the mouse and rat genome sequences and comparisons to the human genome have muddied the waters. With a draft chimpanzee genome sequence available and a high-quality version due soon, the genetic differences between rodents and humans have become more apparent, raising further questions about the validity of mice and rats as human models in drug trials.


Comparing the mouse and human genomes produces some dramatic conclusions. The mouse genome contains 2.5bn DNA letters, around 14 per cent fewer than the human genome, but the human genome contains more repeat sequences than the former. Almost all the 30 000 to 40 000 human and mouse genes are related and have 80 per cent similarity. However, this number could be misleading; some genes are almost identical, while others are barely recognisable as being the same. The genes responsible for smell, immunology and reproduction are most different between humans and mice. These observed differences have allowed scientists to investigate why mice are immune to certain human diseases. They also give a genetic basis to why mice produce many more offspring than humans and have a better sense of smell.

However, the rat genome sequence proved to be even more suprising. Again, almost all rat genes have a related human counterpart, but the human and rat genes are 90 per cent similar. Further, only six of the human genes known to cause disease when defective do not have 1:1 analogues in the rat sequence. Theoretically, this makes the rat a better human disease model.

Similarities and differences
Comparing the three genomes shows around one third of the genome sequences can be lined up for all three species, showing regions that code for important mammalian characteristics. Regions where the sequences line up for the mouse and rat but not for the human genome have also identified where rodent-specific genes are located. Finally, there are clear regions where the genome sequences cannot be aligned. These regions contain species-specific information: regions, in effect, which contain the code to make us human.

With the mouse and rat genomes proving to be so similar to the human genome, it may seem surprising that drug development is hampered by so many failed trials. The problems arise for two reasons. Rodent physiology is quite different to human physiology; the way rodents absorb and excrete drug molecules differs from humans. For example, rats have no gall bladder and can excrete bile very readily. If a drug is excreted via bile, its half-life will be severely reduced in the rat model, possibly suggesting that the drug would be unsuccessful in human trials.

However, the most significant difference in drug molecule activity in humans, rats and mice is the way the chemicals are metabolised. The genomes show humans, mice and rats each produce different amounts and types of detoxifying chemicals. For example, cytochrome P450s (CYPs) have been identified as key substrates for breaking down and removing drugs from the system. In humans, CYP2D6 is an important molecule for metabolising a range of drugs like Prozac and codeine. People deficient in this CYP enzyme are very likely to have adverse drug reactions.

The genetic sequence in humans contains the biological code to produce 57 CYPs; the mouse genome contains the code for 102 and rats have fewer at 53. In addition to the different numbers, rats and mice also produce different types of CYPs to humans. CYP2D6 is our only active 2D enzyme, while in mice there are nine versions; humans have four CYP2c enzymes while mice have 15. These factors suggest that, even using genetically modified mice, which express specific diseases, or inbred rats with engineered mutations, drugs that may combat the disease in humans could be ineffective or toxic in rodents and never make it past the first animal testing phase.

David Nelson, an expert in CYP analysis from the University of Tennessee, Memphis, US, agrees. ’The number of P450 genes in mice is much higher than humans,’ he states. ’It seems unlikely that the substrate specificities and regulation of these additional genes will be the same as in humans.’

So would chimpanzees, our closest ancestral relatives, be better test subjects? The low-quality draft genome sequence was produced in December 2003, with the high-quality draft sequence due soon. Physiologically, the chimpanzee is much closer to humans than rodents are. Comparing the human and chimp genomes has shown our genes to be 98 per cent similar; much higher than the mouse or rat. Theoretically this makes the chimpanzee a better test study for human disease. So far, genetic differences have been identified between the species in the genes responsible for sense of smell, hairiness, digestion and hearing, among others. The latter gives insight into why chimps are difficult to train to understand human speech. Mutations in the human gene alpha tectorin cause poor frequency response in the ear and lead to congenital deafness. Chimp alpha tectorin behaves significantly differently to the human variety, suggesting chimp hearing may not be as well developed as ours.

Large-scale searching for comparisons between chimp and human disease genes is on-going and, as yet, incomplete. It is already known that chimpanzees do not suffer from human diseases such as malaria and AIDS, while they do suffer from diseases that do not affect humans. Scientists are keen to discover the reasons for the species barrier. Nelson has already begun to investigate CYP coding regions in the draft chimpanzee genome and concludes they are almost identical to human CYP coding regions. The number of CYP enzymes produced is also likely to be very similar.

However, even though chimpanzees are likely to be good models for human disease, they are unlikely to become the animal of choice for drug testing and research. ’Chimps are too rare and too precious to use as experimental animals for drug studies,’ says Nelson. Coupled with the public feeling on animal testing, as shown by the demonstrations against a planned primate research facility in Cambridge, UK, it seems realistic to assume chimpanzees will not replace mice as the most common test animal, even if they are more appropriate.

So, where does the information gained from comparative genome analysis leave the world of drug development? Mice are unlikely to be replaced by other animals in testing laboratories because their genes can be easily manipulated and they can be reared and handled simply and cheaply, but comparative genome analysis shows that drug development will continue to be complicated by human/rodent differences.

’An alternative is to "humanise" the mice by replacing the mice CYP gene clusters with human gene clusters,’ says Nelson. This would overcome the problem of mouse CYP enzymes giving different responses to drug molecules than human CYP enzymes, but the physiological differences would still remain. Research into this area is continuing, but this approach may prove to be the best compromise.

There is little doubt that comparative genome analysis is a powerful technique for determining species similarities and differences. With the sequencing of over 200 genomes completed and 1000 on-going, insights into evolution will also be gained. Knowing how our genome differs to others will help to explain what makes us human and give scientists more chance in the fight against disease.


Andrew West is a PhD student at Leicester University, UK, and recipient of the RSC Marriott studentship

The trials of drug development

Using animals to test new drugs is nothing new. In 1885, Pasteur successfully tested vaccines for cholera and rabies on animals and various forms of animal testing have continued ever since. British law currently states that all new drugs must be tested on at least two live mammal species, one of which must be a large non-rodent. With almost every medical treatment used today having been tested on animals, the widespread application of the approach is difficult to overstate.

And so is the success. Animal testing has not only led to licenses being obtained for an enormous number of treatments we take for granted, but it has also given scientists the ability to understand better a range of human diseases including cancer, Down syndrome, diabetes and cystic fibrosis. In 2000 alone, 2.7 million animals were used in research, with the humble mouse being the dominant test animal.

However, it is known that mice are not always the best test models for human diseases. For example, breast cancers are best studied in rats, where hormonal responses are more similar to human responses. It is also well known that results from animal trials are not always reliable indicators for human responses. It is estimated that 70 000 people in the UK suffer from adverse reactions to prescription drugs each year which were not predicted by animal testing. This costs the UK economy millions of pounds in healthcare costs and related expenses. Equally, many accepted treatments with few side effects in humans are ineffective or cause serious side effects in animals; penicillin was originally overlooked because it was inactive in rabbits; aspirin and streptomycin cause birth defects in rats; and ibuprofen causes kidney failure in dogs. With such adverse reactions occurring in test animals, it is possible that successful cures for human diseases have been dismissed before ever reaching a human clinical trial. James Fickett, scientific director of information systems and informatics at AstraZeneca agrees. ’This is certainly possible. In the past, when a drug failed in animal testing, it was very difficult to tell what the failure point might be.’

But Fickett is sure comparative genome analysis will reduce the use of animals in testing and the risk of missed treatments. ’The availability of genomes allows us to test much in the laboratory about the interaction between potential drugs and the specific gene products with which they interact. Then, if and when animal testing is done, we learn much more, and much more quickly.’

Transgenics and gene knockout

Why has the mouse emerged as the test species of choice? Rats were the preferred test animal for over 100 years because of their larger size and better understood physiology. However, during the late 1980s, researchers developed the ability to manipulate mouse genes using knockout technology and transgenics.

Transgenic mice express foreign genes, the DNA for which is introduced into fertilised mouse eggs using microinjection. These mice are born, bred and analysed for the effects of gene overexpression. In gene knockout mice, genes are isolated, inactivated and implanted into mouse stem cells. The cells are then implanted into a female mouse’s womb and allowed to develop. Gene knockout mice are studied for a particular gene’s inactivation effects.

In addition to these two technologies, conditional mouse mutants can be produced in which a gene can be switched off and on at will in different tissues.

All three techniques are designed to produce genetically modified mice that can be used as models for human disease, as well as providing test subjects on which to evaluate new genetic and drug therapies for the disorders.

Such technology is not possible for rats. Rat stem cells are more difficult to culture and the cells fail to develop in the womb. As a result, mice have become more useful than rats or other species for studying the effects of gene defects.