March-April 2013, Volume 10, Issue 2
Embedded in the Red Cell
Published on: March 01, 2013
Professor of Medicine, Harvard Medical School; Physician, Brigham and Women’s Hospital
I have been a science nerd for as long as I can remember. My rite of passage began with my first chemistry set and home laboratory, followed by a series of summer jobs in industrial labs.
During my undergraduate years at Harvard College, I became increasingly intrigued with medical science and obtained an MD in 1961 from the University of Pennsylvania School of Medicine. I then completed a three-year medical residency at New York Hospital, Cornell Medical Center. One of my first admissions was a very gallant and courageous young teenager with Cooley’s anemia (thalassemia). “Alfred” had all of the indications of severe iron overload, including progressive congestive heart failure and endocrine hypofunction that delayed transition into puberty. After he was discharged from the hospital, I saw Alfred in our clinic about once a month over a two-year period, and we became quite close. I will always remember a clinic visit at which he seemed particularly upbeat. He took me aside and said “Dr. Bunn, there is something I want to tell you. On Saturday night I am going out on my first date!” He died about six months later. Alfred and his illness impelled me to go into hematology.
From 1964 through 1967 I was a hematology fellow at the Thorndike laboratory at Boston City Hospital (now known as Boston Medical Center) under the inspiring mentorship of James Jandl, arguably the leading experimental hematologist of that era. Jim’s research focused on the red blood cell and disorders thereof. The intellectual atmosphere at the Thorndike laboratory was transforming, particularly so at a weekly conference in which cases were thoroughly dissected and reassembled by Jim, along with William Castle and Jane DesForges, abetted by younger colleagues, Harry Jacob, Dick Aster, and others who have left a solid imprint on hematology.
After two years in the army at a blood research lab in Fort Knox, KY, I felt I needed an apprenticeship in a strong biochemistry lab before initiating an independent research program. Ruth and Reinhold Benesch had recently reported that the oxygen affinity of hemoglobin in human red cells was tightly regulated by 2,3-diphosphoglygerate (2,3-BPG), an abundant intermediate in the glycolytic pathway. They were aware of experiments I had done at Fort Knox on the impact of falling levels of 2,3-BPG in stored blood, and they gave me the opportunity to join their lab at Columbia University.
Six weeks after my arrival I faced the first and only bona fide crisis in my career. I telephoned Helen Ranney at Albert Einstein College of Medicine and told her that Reinhold had just fired me from his lab, for reasons that I was at a loss to explain. Helen, who at that time was a heavy smoker, replied, “Wait a minute. Let me go get a cigarette. I can see this conversation is going to take a while.” She then arranged for me to join Robin Briehl’s lab at Einstein. While there, I measured the effect of the addition of 2,3-BPG on the oxygen affinity of several human hemoglobins and, after reviewing papers by Max Perutz, made an educated guess regarding sites on deoxyhemoglobin that bind to 2,3-BPG. I communicated these results to Perutz and several months later received an indescribably gratifying handwritten letter telling me that he and Arthur Arnone had x-ray diffraction patterns that confirmed the proposed binding site. Thus began close friendships with Helen and Max, who, with Jim Jandl, were my most influential and generous scientific mentors.
In the 1970s, my lab worked on non-enzymatic glycation of hemoglobin and other proteins. A minor component Hb AIc was known to be elevated in red cells of diabetics. Following earlier work of Bob Bookchin and Paul Gallop at Einstein, we showed that glucose formed a stable adduct with the N-terminal amino of b-globin by a ketoamine linkage. In order to study the biosynthesis of Hb AIc in vivo, I clandestinely infused myself with serum transferrin bound to 59Fe of high specific activity and then monitored the incorporation of radioactivity into the major and minor hemoglobin components. This rather impetuous foray into human experimentation showed that Hb AIc is formed continuously during the red cell’s life span and that the ketoamine linkage is virtually irreversible. Thus, measurement of Hb AIc could be, and indeed has proven, useful in monitoring therapeutic control of hyperglycemia in diabetic patients independent of fluctuations of blood glucose levels.
In 1980, I used my first and only sabbatical leave to go to the National Institutes of Health and work first with Bill Eaton and then with Art Nienhuis. Bill and I addressed a seemingly simple but clinically relevant question: Why is sickle cell trait (AS) benign, whereas sickle cell (SC) patients have significant morbidity? Contrary to an early report, Bill and I showed that polymerization of mixtures of Hbs S and C was identical to mixtures of Hbs S and A. However, we found that two other factors contributed about equally to enhanced sickling in SC patients. First, the level of Hb S in these patients is about 10 percent higher than that in AS individuals because αβS dimers assemble at about the same rate as αβC dimers but more slowly than αβA dimers. In addition, and of equal importance, polymer formation is favored in SC red cells because Hb C induces water loss and thus higher intracellular hemoglobin concentration.
My six months in Art’s lab provided me with sorely needed hands-on tutelage in molecular genetic technology and enabled my lab to switch its focus from hemoglobin to erythropoietin. This transition was greatly facilitated by the creative input of a research fellow Mark Goldberg, who, during college and medical school, had worked with me on sickle hemoglobin. After an exhaustive search, Mark found two human hepatoma cell lines that produce erythropoietin in response to hypoxia. These cells enabled us and others to identify key elements in the Epo gene that are important in hypoxic induction. Jean-Paul Boissel and I prepared a large number of site-directed mutants of erythropoietin that helped to confirm its three-dimensional structure and also to determine the sites that bind to the erythropoietin receptor.
I have also had the opportunity to oversee some timely and fulfilling clinical research projects including one of the early trials of hydroxyurea therapy for SC patients and the use of subcutaneous deferroxamine for treatment of iron overload in transfusion-dependent patients with myelodysplasia.
During my tenure at Brigham and Women’s Hospital, I have tried to fulfill a meaningful role in patient care, a task made a lot easier and more fun by having the opportunity to work closely with remarkable colleagues. My predecessor as chief of hematology was William Moloney, a master clinician with uncanny wisdom reinforced by boundless energy and wit. My successors, Bob Handin and Nancy Berliner are also superb clinicians who have not only taught me a lot about hematology, but have also been supportive and tolerant of my idiosyncrasies, geriatric and otherwise. I never found it easy to remain viable in competitive areas of research while hanging on to and hopefully enhancing my clinical skills. However, this ongoing challenge has been greatly ameliorated by exposure to two generations of extraordinarily bright and effective fellows, house staff, and students at the Brigham and Harvard Medical School.
I am also indebted to the American Society of Hematology. I was fortunate to have helped in the leadership of this wonderful organization, including a role in enabling ASH to own and publish Blood and a 10-year stint as an associate editor. For a while, I was “impresario” of the musical events that were previously such a haven of civility and repose at our annual meetings. On one blissful occasion, prior to an ASH concert in San Francisco, I had the opportunity to play the piano in a movement of a Schubert trio with Yo Yo Ma and violinist Lynn Chang! The Society fosters a remarkable climate of collegiality and cooperation that has been an enormous boon to hematology in an era when our specialty has faced a variety of challenges that threaten its livelihood.
Thoughts From a Former Protégé
Mark A. Goldberg, MD
Senior Vice President of Medical & Regulatory Affairs, Synageva Biopharma Corp. Clinical Associate Professor of Medicine, Harvard Medical School
Webster’s Dictionary defines a mentor as “a trusted counselor or guide.” I would go a step further. To me a true mentor is someone whom you can rely upon to give you sound, experienced direction that, above all else, is in your own best interest, even if that conflicts with the mentor’s best interest. By this definition, true mentors are a very rare breed. In Frank Bunn, I have been fortunate enough to have been mentored by one of the best of this rare breed for more than 39 years.
I first met Frank in the fall of 1973. I was a college sophomore with a little lab experience that I gained during high school, and Frank was a young assistant professor of medicine at Peter Bent Brigham Hospital and Harvard Medical School. After interviewing with Frank, he offered me a position in his laboratory. Little did I know how dramatically and positively Frank would impact my personal and professional life.
I worked in Frank’s lab throughout college and during medical school and then did my postdoctoral research with him. Under Frank’s mentorship, I learned a tremendous amount about science, hematology, and life. During that time he taught me how to rigorously approach scientific problems and how to design and conduct well-controlled experiments. I learned about the molecular basis of sickle cell disease and how it leads to many of the clinical manifestations of the disease. We studied the regulation of erythropoietin gene expression by hypoxia and made a number of interesting observations.
After I had started my own lab as an independent investigator, Frank remained very supportive. I still spoke often with him. Once when I was preparing a manuscript for publication, I discussed with Frank having him as a co-author. Frank felt it was important for me to publish independently of him and suggested that he not be a co-author.
Throughout my formative years as a hematologist, Frank always made sure to introduce me to leaders in the field. Shortly after I received my first R01 award, Frank told me that Ernie Beutler wanted to have lunch with me at ASH to discuss recruiting me to Scripps. During lunch, Ernie shared with me that when he told Frank about his desire to recruit me, Frank appeared as if he was giving away his first born. Nonetheless, Frank made the introduction because he wanted me to have every opportunity.
For close to four decades I have had the privilege of seeing firsthand the hard work and innovative thinking that allowed Frank to make very significant contributions to our basic understanding of hemoglobin structure and function, as well as the regulation of erythropoiesis, and to translate these findings to clinical applications that have helped patients with diabetes mellitus and sickle cell anemia. I have also seen Frank’s commitment to teaching, as he has assumed a leadership role in teaching hematology to generations of Harvard medical students.
My career has evolved in ways that I had never imagined. While still maintaining my academic appointment at Harvard and the Brigham, I have spent most of the past 15 years working in the biotechnology industry. I still frequently seek out Frank for his advice and opinions. We get together regularly for lunch. I always look forward to these dates. We still talk about science and medicine but much more about our lives and our families. I have been extremely fortunate to have such a caring, thoughtful mentor. I hope that I can pay forward what I have learned from Frank about the meaning and value of true mentorship. I hope I can be like Frank.
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