杏吧原创

WHEN a 25-year-old man was brought to the King Edward Memorial Hospital in Mumbai, India, his outlook was bleak. Sanjay^* had severe aplastic anaemia, a condition where the bone marrow stops making enough red blood cells, immune cells and platelets. He needed five blood transfusions a month just to stay alive, and even so, he was bleeding from his gums and eyes, and could barely get out of bed.

That was July. Barely three months later, Sanjay is in rude health, up and playing cricket with his friends. When New 杏吧原创 interviewed him last month, Sanjay had no signs of bleeding or jaundice. He was bright, lively and full of smiles.

Sanjay鈥檚 reversal of fortune is apparently thanks to a controversial new stem-cell treatment from a little-known British biotech firm called TriStem. The firm鈥檚 scientists make the provocative claim that they can make customised stem cell therapies simply by taking a sample of someone鈥檚 blood and treating their immune cells with a special antibody for a few hours.

And while most mainstream stem cell research is still in the Petri dish, TriStem this week announced that the first four patients to receive its treatment have made dramatic recoveries from their aplastic anaemia within days. What鈥檚 more, the firm says its technology holds similar promise for the wide range of debilitating diseases that mainstream stem cell researchers are tackling, such as heart failure, spinal cord injury and type 1 diabetes.

If previous experience is anything to go by, these claims are likely to draw guffaws of disbelief. But while the latest results fall short of definitive proof that the technique works, they are certainly intriguing. 鈥淚f it鈥檚 true, it鈥檚 a fundamental rewrite of the textbooks,鈥� says Richard Boyd, a stem cell researcher based at Monash University in Melbourne who has reviewed their data first-hand. It would be, he says, 鈥淣obel prize-winning stuff鈥�.

Stem cells are one of the hottest research areas in medicine right now because they offer the possibility of regenerating seemingly any part of the body (see 鈥淲hat are stem cells?鈥�). But while millions of pounds a year are being ploughed into the field, research is at a very early stage. There is wide disagreement about which cell surface markers identify the different stem cell types, and which tissues they can develop into. A handful of human studies are taking place 鈥� mainly involving bone marrow stem cells, whose normal role is to produce the cells of the blood system 鈥� but the vast majority are at the animal research stage, or are looking at cultured cells in the lab.

That is what makes TriStem鈥檚 claims so astonishing. If they are valid, the firm鈥檚 scientists could have made a breakthrough comparable to Alexander Fleming鈥檚 discovery of penicillin 76 years ago.

Like penicillin, TriStem鈥檚 technology was discovered by accident. In 1990, Ilham Abuljadayel was a consultant immunologist at a military hospital in Jeddah, Saudi Arabia. In her spare time she started carrying out some leukaemia research.

Leukaemia occurs when someone鈥檚 immune cells mutate and turn cancerous, multiplying out of control. Abuljadayel knew that the cancerous cells bristle with unusually high numbers of a surface molecule called MHC II. An antibody called CR3/43 that binds to MHC II was commercially available (from a Danish firm called DakoCytomation). Abuljadayel started investigating if leukaemia cells could be killed by exposing them to a solution of CR3/43 along with certain immune molecules called complement, which destroy anything bound to antibodies.

The approach had some success, but one day Abuljadayel accidentally omitted the complement from a batch of solution. Something strange happened. Instead of dying, the dark leukaemia cells visibly changed, becoming transparent and apparently flourishing. Frustrated, Abuljadayel threw away the culture and repeated the experiment with more of the same solution. Again she saw the same result. After several further attempts it occurred to Abuljadayel that the transparent cells had the distinct appearance of bone marrow stem cells.

Biological heresy

Abuljadayel confirmed her hunch with some basic tests and discovered her mistake with the complement. She began wondering if the antibody binding to MHC II had somehow reversed the process of specialisation or 鈥渄ifferentiation鈥� that occurs during normal development in the womb, when embryonic stem cells develop into all the tissues of the body. The leukaemia cells seemed somehow to have 鈥渞etrodifferentiated鈥� 鈥� a biological heresy.

She found that some of the cells looked like various types of blood cells that had never been present in the original sample. Some of the newly formed bone marrow stem cells could have subsequently re-specialised into the other blood cells, she concluded. 鈥淭he entire [blood] system was created from these leukaemic cells,鈥� she told New 杏吧原创. 鈥淎t first, I thought I was going nuts. I repeated it again and again.鈥� Encouragingly, Abuljadayel also found that the antibody had the same effect on normal non-leukaemic immune cells, which also have MHC II on their surface, albeit less.

Abuljadayel tried to get her work published, but without success. 鈥淚 wrote up a lot of pieces, and it was rejected every time,鈥� she recalls. At the time, retrodifferentiation of cells was considered plain impossible. This was before the arrival of Dolly the cloned sheep in 1997, who was also created by regressing an adult cell nucleus, albeit by a different method.

Abuljadayel and her husband took out patents on the technology in 1994, and later founded TriStem to commercialise it. Funded by private British backers, they refined the technique and discovered that it worked even better with a special purified form of the CR3/43 antibody, also made by DakoCytomation.

As if the initial work weren鈥檛 controversial enough, in the late 1990s the firm dropped a further bombshell. The antibody could make immune cells retrodifferentiate even further back than bone marrow stem cells, back to a state with the versatility of embryonic stem cells, they claimed. And by subsequently exposing these cells to the same growth factors that other scientists were already using to manipulate conventional embryonic stem cells, Abuljadayel said she could produce in the lab virtually any tissue type 鈥� neurons, muscle, liver, pancreas, heart 鈥� you name it.

If these claims are true, it could give doctors the ability to treat patients with tailor-made genetically matched cells, created from a mere blood sample. It would also sidestep the ethical minefield surrounding the use of embryonic stem cells for this purpose.

In contrast to the accepted mode of disseminating science, however, little of this work appeared in peer-reviewed journals. The first most people heard of it was when The Times newspaper in London published an article about it on 15 January 2001.

It was only last year that TriStem published two papers in a peer-reviewed journal, Current Medical Research and Opinion. The first described how they used the antibody and growth factors to generate several cell types in culture, including neurons and heart muscle cells (vol 19, p 355). The second showed that if human immune cells were treated with the antibody and injected into immunosuppressed mice, they entered the animals鈥� bone marrow and generated nearly all the different types of cell present in human blood (vol 20 p 87).

Importantly for TriStem鈥檚 credibility, the second study was a collaboration with a respected team of stem cell researchers led by Tim McCaffrey at George Washington University in Washington DC. Abuljadayel had got McCaffrey on board through an eye-catching demonstration. When heart muscle cells are grown in culture, they spontaneously start rhythmically contracting, like a miniature simulation of the full-sized organ. Abuljadayel had taken a blood sample from McCaffrey himself and incubated his cells with the antibody and the growth factors that make heart muscle cells. A few hours later, she invited him to look down the microscope. There before his very eyes, what had once been his blood cells were now beating. 鈥淚t鈥檚 stunning,鈥� he told New 杏吧原创 at the time (29 November 2003, p 6).

Many scientists were still sceptical. Some disagreed with the team鈥檚 choice of cell surface markers to signify bone marrow stem cells. Alexander Medvinsky of the Institute for Stem Cell Research in Edinburgh suggested that the antibody might just be killing most of the immune cells to leave only stem cells, which then multiplied. (A few bone marrow stem cells are present in blood.) But to this McCaffrey countered that tests from the original research showed that about 90 per cent of the original cells survived.

Since then McCaffrey has continued to work on the heart cells that Abuljadayel created. McCaffrey says he has already had some success using them to treat an animal model of heart disease, although he won鈥檛 give details until a paper has been published.

But TriStem鈥檚 most compelling results have come from its latest work, in aplastic anaemia. This condition, which is more common in developing countries than in the west, occurs when bone marrow stem cells are damaged and stop generating certain blood cells and platelets. The cause is unknown, but suspected culprits include malnutrition, a viral infection, or exposure to toxic chemicals.

As the condition progresses, patients become weak from anaemia and suffer infections, and their blood fails to clot properly. They need regular blood transfusions just to stay alive, and the only long-term hope is a bone marrow transplant from a genetically matched donor. Few in poor countries have this option, and even in the west, suitable matches may not be found. If they are, the problems don鈥檛 stop there: patients have to take immunosuppressive drugs for the rest of their lives, and the transplant may still be rejected.

TriStem formed a collaboration with the Indian Council of Medical Research, based in New Delhi, Dipika Mohanty, director of the Indian Institute of Immunohaematology, and an Indian blood products firm called GG HaemHealth. The initial plan was to harvest immune cells from the blood of a healthy sibling, treat these with the antibody and then inject patients with the resulting bone marrow stem cells. But Nirmal Ganguly, head of the ICMR, convinced the team it would be safer to use the antibody on what few immune cells the patients had left, rather than involving donors.

After getting approval from the hospital鈥檚 ethics committee, the Indian collaborators selected the first patients for TriStem to treat, in July. They certainly chose difficult cases. 鈥淪ome were on death row,鈥� says Abuljadayel. Sanjay went first. His cells were exposed to the antibody for about two-and-a-half hours, cleaned, concentrated, and re-injected.

Sanjay recalls what happened: 鈥淚 woke up the next day and I felt so strong I wanted to get up and run out of the hospital.鈥� Bored, he started asking to go home. A biopsy of his bone marrow two weeks on showed that what had been a barren wasteland was now teeming with cells. Analyses showed his blood system seemed to totally recover within about a week; in comparison, it takes about three months for a patient to recover after a bone marrow transplant. In the bleak world of aplastic anaemia, it seemed like a miracle.

First step

The researchers performed the procedure on three more patients. One was particularly ill, bleeding from every orifice and needing 12 transfusions a month. All showed similarly spectacular recoveries, with their blood cells reaching desired levels within at most, 20 days of the treatment. They鈥檝e collectively avoided some 60 transfusions since their treatment. 鈥淭hey鈥檙e leading a normal life, going out with friends and enjoying things,鈥� says Abuljadayel.

As New 杏吧原创 went to press, TriStem was preparing to announce its results on its website this week. Unsurprisingly, mainstream stem cell researchers are sceptical. For Evan Snyder, a leading stem cell researcher at the Burnham Institute in La Jolla, California, the change from immune cell to stem cell happens unfeasibly fast. 鈥淭he time frame looks suspect to me,鈥� he says. 鈥淐ells can鈥檛 be born that quickly.鈥�

So how likely is it that TriStem is taking everyone for a ride? Or could they simply be misguided about what they think they鈥檙e seeing? Not according to Richard Boyd, an independent stem cell researcher from Monash University in Melbourne, who witnessed the procedure in Mumbai from start to finish. 鈥淭hey are genuine people,鈥� he says. 鈥淲e saw when the blood came out of the patient, and after it had been incubated with the antibody there was a visible difference. After six days, this guy had platelets and neutrophils he never had before. In all four patients, they鈥檝e had an effect.鈥�

Assuming Boyd is right, this is still only a first step for TriStem. If the firm is to get the antibody treatment approved as a therapy for aplastic anaemia, regulatory authorities will doubtless want to see decent-sized placebo-controlled trials carried out. The firm is consulting Quintiles, a major contract research organisation, about how to proceed.

Controlled trials are considered essential to prove beyond doubt the technique works and that it does not do more harm than good. The antibody-treated cells could in theory turn cancerous, for example, and cause leukaemia, says Boyd (although aplastic anaemia patients sometimes develop leukaemia anyway). Equally theoretically the modified cells could mount an immune response against the patients, although Boyd thinks the risk of either happening is 鈥渇airly minimal鈥�, and would apply equally to other stem cell therapies.

And it鈥檚 not yet clear just how retrodifferentiated the modified cells are. It is unknown if they are true bone marrow stem cells known as haematopoietic stem cells, which can self-regenerate indefinitely and so could be a permanent cure. Boyd鈥檚 hunch is that they could be less retrodifferentiated 鈥減rogenitor鈥� stem cells which, while capable of producing all the cells of the blood system, cannot self-regenerate. In that case, the antibody treatment might need to be repeated every few months, although this would still be a major advance over current therapies.

Abuljadayel says as much as 20 per cent of the modified cells had surface markers that suggest they were haematopoietic. She says the fact that the first two patients have now reached three months without a blood transfusion suggests this was indeed the case. Boyd intends to investigate this further with TriStem by genetically profiling the treated cells.

More research is also needed to find out how the antibody technique works, assuming it does. These days the idea of retrodifferentiation is no longer seen as quite so fanciful 鈥� since Dolly, a menagerie of cloned animals have been produced by various methods. But just how an antibody to the MHC II cell surface molecule could have such a remarkable effect is still a mystery. MHC II鈥檚 primary function is to allow immune cells to distinguish between harmful pathogens and the body鈥檚 own cells.

TriStem has still not convinced many mainstream scientists of its most provocative claim, that it can retrodifferentiate blood cells back to a state with the versatility of embryonic stem cells, and so create the other kinds of tissues such as neurons and heart cells. The firm plans to next try its treatment on people with spinal cord injury, heart disease and leukaemia but it is far too soon for patients with these conditions to get their hopes up. Even if TriStem succeeds in these other fields, it can sometimes take decades to translate advances in basic science into treatments used in the clinic.

But at the very least the field of aplastic anaemia may get a much-needed shot in the arm. Abuljadayel believes she has already helped four people beyond anyone鈥檚 expectations, and that will be tricky for sceptics to ignore. 鈥淚 don鈥檛 know if they鈥檙e cured,鈥� she says. 鈥淏ut if they hadn鈥檛 been treated, who knows how much longer they would have survived?鈥�

And Abuljadayel is already thinking beyond anaemia. 鈥淚t鈥檚 just a start,鈥� she says.

*Not his real name