The Nobel Prize in Chemistry 1993

Michael Smith

Protein Testing in Lymphoproliferative Disorders

Serum protein electrophoresis with or without certain additional tests is required on all new patients with multiple myeloma or other plasma cell disorders, malignant lymphoma and all patients with chronic B or T lymphoid leukemias. These tests are not required in new patients with a diagnosis of Hodgkin's disease. The terminology of protein testing and recommendations for specific tests are as follows:

1. Serum Protein Electrophoresis (SPE) SPE is performed by cellulose acetate or high resolution agarose gel electrophoresis. The latter technique is very sensitive and useful for the detection of small monoclonal paraprotein bands. In the majority of patients with myeloma, SPE is useful for diagnosis usually demonstrating a monoclonal protein, associated hypogammaglobulinemia or both. For long-term monitoring of patients with clinically significant monoclonal proteins, quantitation of the paraprotein by SPE is preferred to other available tests. It more accurately estimates the actual amount of protein present than quantitative immunoglobulin studies.

2. Immunofixation (IFE) Once a paraprotein band is identified by SPE, the type of the paraprotein band is determined by IFE. This technique is very sensitive and has, for the most part, replaced immunoelectrophoresis. In any disorder characterized by a paraprotein, this test is usually only required at diagnosis to establish the nature of the paraprotein. If a monoclonal protein is identified in the urine (Bence-Jones protein) by urine protein electrophoresis (UPE), the identity of the paraprotein is also determined by IFE.

3. Quantitative Immunoglobulin Levels The specific amounts of individual immunoglobulins by class (IgG, IgM, IgA) can be determined by several methodologies. Two of the more common methods are radial immune diffuse and nephelometry. These measurements are of utility in patients with immune deficiency, however, they are of minimal value in patients with neoplastic lymphoproliferative disorders with a paraprotein. Occasionally it is useful to determine that the other classes of immunoglobulin are decreased in a patient with a paraprotein.

4. Urine-Protein Electrophoresis (upe) A 24 hour urine sample is usually required in patients with a neoplastic paraprotein. The urine sample is concentrated and then fractionated by protein electrophoresis. The amount of protein is quantitated and the pattern is examined to determine if a paraprotein band is present. If a band is detected the specific type is identified by IFE. Older qualitative tests for presence of Bence-Jones protein are no longer useful.

I was born on April 26th, 1932 at 65 St. Heliers Road, South Shore, Blackpool, England in the house of my maternal grandmother, Mary Martha Armstead, having been delivered by the District Nurse, Ms. Parkinson, a lady who I can remember from my infant and juvenile days in her uniform and navy blue raincoat on her bicycle doing her rounds and visiting schools for health inspections. My parents, Mary Agnes Smith and Rowland Smith, both had to work since their early teens, she in the holiday boarding house of her mother and he in his father's market garden in Marton Moss, a village on the south side of Blackpool, just north of Saint Anne's-on-Sea. I went to the local school, Marton Moss Church of England School for 6 years from the age of 5. My mother attended the local church, Saint Nicolas, and consequently I attended that church and its Sunday School. My only prizes from the Sunday School were "for attendance", so I presume my atheism, which developed when I left home to attend university, although latent, was discernible.

During my last year at elementary school, 1943, I sat for the "Elevenplus" examination which was used in the English schools in those days. In principle, of course, it was an invidious system designed to identify the approximately 20% of the school population that would be offered an academic education and the 80% who would be obligated to take a secondary education that terminated with no further academic options at age 15 (of course, there was the alternative of private schooling, but that was not an option if you were the child of poor parents, as was I). I was lucky enough to obtain a scholarship to the local private school, Arnold School. I did not, at the time, consider this to be luck. I did not want to go to Arnold School because the pupils were considered to be snobs and I thought that I would be ostracized by my friends in Marton Moss. Luckily, my mother insisted, and I went to Arnold School. I cannot say it was the happiest time in my life (I was no good at sports, and proficiency in sport is important in private school life. And I hated the war-time meals that were provided at lunch, as well as the prefect who insisted that I eat the awful food). But the schooling was first-rate, and in this I flourished, although not equally well in all subjects. Clearly, science was my metier, and I was lucky to have a chemistry teacher, Sidney Law, who stimulated my interest in chemistry and who took a personal interest in me (he told me I should read a better newspaper than the one to which my parents subscribed and, as a consequence, I became a life-long reader of the Manchester Guardian. That, in turn, stimulated me to become a reader of the New Yorker as soon as I came to North America, another life-long addiction).

The seven years from 1943 to 1950 were also a time when I became a boy scout. That was a piece of luck. The headmaster at Arnold School, Mr. Holdgate, at the end of my first term, sent me to a dentist, Mr. Paterson, in the hope that he could correct my protruding front teeth, about which I had been teased by my schoolmates. Mr. Paterson did not correct the problem with my teeth but he did introduce me to a wonderful scoutmaster, Mr. Barnes, who inducted me into the happy world of camping and outdoorsmanship which provided me with enjoyment and vacations throughout my secondary school years and right up to the present. An enjoyment that explains why I have a particular delight in living amid the rugged outdoors and beauty of British Columbia.

The second World War impinged on the lives of many of us who were alive at the time. Blackpool, as it turned out, was a very safe place, being in the northwest of England and distant from the targets for German bombing. The large number of hotels and boarding houses in this seaside resort were used to house military trainees, mainly for the airforce. And my father, working his father's market garden, grew primarily food crops rather than his preference, chrysanthemums. Occasionally, bombers, presumably diverted from their primary targets of Manchester and Liverpool, would try to bomb the new factory behind our house that produced Wellington bombers. Usually, they hit the market gardeners' greenhouses which showed up better at night. And I remember one night, alone in the house with my baby brother Robin, when a stick of bombs fell on either side of the house.

I was not proficient in Latin and so was not able to go to Oxford or Cambridge. However, I did enter the first-rate chemistry honours program at the University of Manchester in 1950, where the professors were E.R.H. Jones and M.G. Evans, and graduated in 1953, with the financial support of a Blackpool Education Committee Scholarship. I had hoped to get a firstclass degree, but only got a 2(i)! I was very disappointed. However, I still was able to obtain a State Scholarship which supported me throughout my graduate studies until I finished my Ph.D. degree in 1956. My supervisor was H.B. Henbest. He was an outstanding young organic chemist, and I was glad to have him as a supervisor of my work on cyclohexane diols. However, we did not have a particularly warm relationship. I was socially shy and moody and was probably quite hard to understand.

The last year of our graduate studies saw me and my classmates writing to various American professors seeking post-doctoral fellowships. I had no luck in obtaining my desire of a fellowship on the west coast of the United States, but I heard, in the summer of 1956, that a young scientist in Vancouver, Canada, Gobind Khorana, might have a fellowship to work on the synthesis of biologically important organo-phosphates. While I knew this kind of chemistry was much more difficult than the cyclohexane stereochemistry in which I was trained, I wrote to him and was awarded a fellowship after an interview in London with the Director of the British Columbia Research Council, Dr. G.M. Shrum.

I arrived in Vancouver in September 1956. My first project was to develop a general, efficient procedure for the chemical synthesis of nucleoside-5' triphosphates based on the synthesis of ATP by Khorana in 1954. This study led to more extensive investigations of the reactions of carbodumides with acids, including phosphoric acid esters and to a general procedure for the preparation of nucleoside-3',5' cyclic phosphates, a class of compounds whose existence and great biological significance had only recently been discovered. One particular pleasure of that period was the development of the methoxyl-trityl family of protecting groups for nucleoside-5'-hydroxyl groups (one synthesis of trimethoxytritanol erupted and left a large orange stain on the laboratory ceiling); this class of protecting group is still in use in modern automated syntheses of DNA and RNA fragments.

In 1960, the Khorana group, including myself, newly married (I have three children, Tom, Ian and Wendy. My wife Helen and I separated in early 1983), moved to the Institute for Enzyme Research at the University of Wisconsin. There I worked on the synthesis of ribo-oligonucleotides, that most challenging of chemical problems for a nucleic acid chemist. Early in 1961, I began to realize that it was time to move on. Helen and I wanted to return to the West Coast of North America, and I accepted a position with the Fisheries Research Board of Canada Laboratory in Vancouver. I enjoyed my time there because of the opportunity it presented to learn about marine biology and I was able to sustain my interest in nucleic acid chemistry because of the award of a U.S. National Institutes of Health Grant, which led to a new synthetic method for nucleoside-3',5' cyclic phosphates. However, the atmosphere of the laboratory, although based on the campus of the University of British Columbia, was not really conducive to, or supportive of, academic research. Hence, in 1966, I was very glad that Dr. Marvin Darrach, then Head of the Department of Biochemistry, offered to nominate me for the position of Medical Research Associate of the Medical Research Council of Canada. This award, which provided salary support, allowed me to become a faculty member of the Department, my academic home ever since, except for sabbaticals at Rockefeller University, the Laboratory of Molecular Biology of the Medical Research Council in Cambridge and Yale University. The Council also has provided research grant support throughout my academic career.

In 1981, Ben Hall and Earl Davie, of the University of Washington, invited me to be a scientific cofounder of a new biotechnology company, Zymos, which was funded by the Seattle venture capital group, Cable and Howse. One of the first contractors was the Danish pharmaceutical company, Novo, who asked Zymos to develop a process for producing human insulin in yeast. After a considerable cooperative effort by Zymos and Novo researchers a successful process was developed. In 1988, the pharmaccutical company, now Novo-Nordisk, purchased outright the biotechnology company, now named ZymoGenetics. I am pleased that, although I no longer have any involvement, ZymoGenetics has subsequently expanded and has continued research on a wide variety of potential protein pharmaceuticals.