In February 1997, world famous tenor José Carreras sang happy birthday to an Australian scientist, Professor Donald Metcalf. It was quite a birthday party, and there was good cause to celebrate. Ten years earlier, Carreras had been diagnosed with leukaemia and was fighting for his life. His recovery was due, in part at least, to painstaking research conducted by Professor Metcalf and his colleagues into colony stimulating factors. The research was carried out at the Walter and Eliza Hall Institute in Melbourne. Colony stimulating factors regulate white blood cell production White blood cells form the basis of the body's immune system, guarding against attack by viruses and other microorganisms. Colony stimulating factors are proteins that help regulate the production of white blood cells. These cells are short-lived, so we need to produce a lot of them: healthy adults produce about 200 million new white blood cells every hour, mostly in the bone marrow. White blood cells are not the only cells produced in the bone marrow. Red blood cells (also called erythrocytes) and platelets (which aid blood clotting) are also produced there in large numbers. Astonishingly, all these cells appear to derive from a single type of master (or stem) cell, called the haematopoietic (blood-forming) cell. For decades, scientists thought that some sort of mechanism must exist to regulate the production of white blood cells from these haematopoietic cells. But, despite many attempts to discover such a mechanism, its eventual detection was an accident. A lucky breakthrough In the mid 1960s, scientists in Israel and Australia independently attempted to grow mouse leukaemia cells in nutrient agar plates. Although unable to do this, they observed the spectacular growth of healthy white blood cells in clusters or 'colonies' around other tissue fragments in the agar. Further experiments showed that the number and size of the white blood cell colonies were dependent on adding other cells or tissue fragments to the cultures. Scientists speculated that some factor contained in this material was stimulating haematopoietic cells to divide and form these colonies. This still unidentified factor was dubbed a 'colony stimulating factor' (CSF). Searching for CSFs The circumstantial evidence for the existence of such a factor grew, prompting Metcalf's team to attempt to extract or purify it from tissues. This was not an easy task: while a colony stimulating factor appeared to be present in all organs of the body, it was there only in very small quantities. It was a bit like searching for a needle in a haystack, except that the shape and size of the 'needle' was unknown. The project to purify CSF lasted 15 years. As the years went by, new technology became available to help in the task. It was also realised that more than one CSF existed: eventually, the project identified four distinct CSFs, each responsible for stimulating the growth of different kinds of white blood cell. The nature of CSFs was also explored. Scientists determined that they were glycoproteins, molecules that combine a protein and a large sugar molecule (polysaccharide). Studies revealed that CSFs stimulated the division of immature white blood cells and were essential for the continued development of such cells. CSFs were also shown to influence the ability of certain white blood cells to kill microorganisms. Little by little By 1984, scientists had purified several human and mouse CSFs in small quantities. But they were gloomy. The hard work of the past decade, in several laboratories, had failed to produce enough purified CSF to test a single laboratory mouse. In fact, calculations showed that at the current rate of purification it would take 250 years to produce enough of the substance to treat a single human patient for 2 weeks. The race to mass-produce A new way was needed, and it was at hand. Molecular biologists were already experimenting with techniques such as gene cloning and the artificial production of enzymes, hormones and other proteins using genetic techniques. The biologists thought that by isolating the genes encoding the various CSFs and inserting them into yeast cells, bacterial cells or mammal cells, it should be possible to produce CSFs in much larger quantities. The potential of mass-produced CSFs to aid the treatment of disease attracted the interest of a number of commercial enterprises. In the years 1984 to 1987 they raced each other to discover - and patent – the genes responsible for CSF production. Success was almost immediate, and mass production of CSFs followed soon after. A big day in the lab Despite this rush of activity, the value of CSFs in the treatment of disease was still untested. Professor Metcalf and his team, now armed with relatively large quantities of several CSFs, set out to see what effect the injection of one particular CSF might have on mice. After a week of injections, white cell counts in blood samples taken from the mice showed virtually no change. According to Professor Metcalf, 'for an hour gloom reigned in the laboratory'. The mood changed dramatically with the next sample, taken from the abdominal cavity: the effect was obvious, even to the naked eye. The fluid was milky, due to the presence of a large number of white blood cells. This contrasted with the clear fluid obtained from mice that were not injected with the CSF. Dejection in the lab turned to euphoria. The evidence seemed clear: the injection of CSFs resulted in increased production of white blood cells. The role of CSFs in leukaemia treatment So far this experiment had only been performed on mice. A far more important task was to test the treatment on humans. José Carreras was one of the first people to be treated with a CSF. He suffered from acute myelogenic leukaemia, a cancer of the blood that is usually fatal within months if untreated. Unlike other cancers, leukaemia doesn't produce tumours but instead causes the rampant overproduction of cancerous white blood cells. Such cells interfere with vital organ functions, including the production of healthy red and white blood cells and platelets. Often, the only treatment for acute leukaemia is a bone marrow transplant, which usually proceeds in three stages. The first involves intensive chemotherapy or other treatment to bring the patient's cancerous white blood cell count under control. In the second stage, the patient's bone marrow is first destroyed by intensive chemotherapy to avoid rejection of the new marrow and a new marrow from a compatible donor is then inserted. The third stage, recovery, is often the most dangerous. Until the donor marrow cells start producing new blood, the patient is left with virtually no immune system. This makes infection very likely. It is at this stage that CSFs play their part. Their injection into a patient has a dramatic effect on the number of white blood cells produced in the early days after a bone marrow transplant, significantly increasing the patient's chance of survival. José Carreras recovered completely, and he now sings the praises of the treatment. Many other bone marrow transplant recipients have since also benefited from treatment with CSFs. Other roles for CSFs? CSFs have a potential role to play in the treatment of other cancers. Infection is the most common cause of death among cancer patients undergoing chemotherapy because the production of white blood cells is affected. CSFs can be injected into such patients after chemotherapy, increasing the speed at which white blood cell counts return to normal. This also allows the possibility of increasing the chemotherapy dosage, since loss of immunity may not be such a significant problem. Other possible uses for CSFs continue to be explored by scientists around the world, including in Australia, where it all began.
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Posted March 1998.
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This topic is sponsored by The Walter and Eliza Hall Institute of Medical Research.
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