Tee SoS ee Se Tee RT TSaST EY edly. Glactl eo, Jaffe, Keun aye ed ve sreoyuen, ae ch Raw Washi Na ACADEMIC MEDICINE PRESENT AND FUTURE Editors John Z. Bowers, M.D. Edith E. King ROCKEFELLER ARCHIVE CENTER CONFERENCE Pocantico Hills North Tarrytown, New York 25-27 May 1982 CYCLES AND FASHIONS IN BIOMEDICAL RESEARCH JOSHUA LEDERBERG My task was to collect some of the threads comprising the fabric of fundamental biology and to comment on the health and medical appli- cations thereof. As shorthand for that conception, the elaboration of biology and pathology from first principles of chemical and physical structure, I will caption it a reductionist or reductive model. The starting point of my own thought was very well stated by Drs. Kennedy and Lehninger, who talked about the promissory notes that reductive biology had been tendering for a number of years. Dr. Kennedy quoted Dr. Charles Huggins: “Whose lives have been saved by a Warburg apparatus?” J suspect that is not such a difficult question to answer. My variant ts, “How many lives have been saved in the last twenty years by the ‘double helix’?”—an expression that stands as proxy for all of modern reductive biology. In 1944 Avery, MacLeod, and McCarty reintroduced DNA to the Josuua LEDERBERG graduated from Columbia College with a B.A. degree in 1944. He enrolled in the university's College of Physicians and Surgeons while he worked as a research assistant in genetics. After two years he terminated his medical studies and moved to Yale University where he received his Ph.D. in senctics in 1947. Dr. Lederberg spent the next ten years in the Department of Genetics, College of Agriculture, at the University of Wisconsin, where, in 1957, he created and became Chairman of the Department of Genetics at the university's medical school. Dr. Lederberg moved to Stanford University School of Medicine in 1959 as Professor of Genetics, as well as of Biology and Computer Science, and Chairman of the Department of Genetics. In 1978 he became President’ of the Rockefeller University. His fields of interest include genetics, chemistry, and evolution of unicellular organisms and of man, computer models of scientific reasoning: exobiology and the origin of life: and applications of scientific understanding to public health and policy. In 1958 Dr. Lederberg shared the Nobel Prize for studies on the organization of genetic material in bacteria. 202 J. Lederberg CYCLES IN BIOMEDICAL RESEARCH biologists’ consciousness. This development stood against the presumption of the prior two decades that proteins were everything: they were enzymes, and they were sources of such exquisite specificity in every other realm, why not in the genetic material as well? But as is well known, the experiments of these investigators gave the first and eventually irrefutable, direct evidence that genctic specificity resided in the chemical structure of DNA. In the brief interval from 1944 to the beautiful elaboration of the structure of the DNA molecule, the double helix, by Watson and Crick in 1953, thinking and experiments in biology were unassailably revolu- tionized. Little biological research today is not decply informed by these conceptions. Nevertheless, until just now, one might have sought in vain for impor- tant public health or specific therapeutic applications of that knowledge. As a geneticist, I would be the first to recall many important applications of chromosome and cell biology, ¢.g., the delineation of genetic syndromes and the further illumination of pathogenetic processes. Starting with Garrod’s insights, the development of medical genetics followed soon upon the rediscovery of Mendelism in 1900. It is all the more paradoxical that hardly anybody’s health for twenty-five years after 1953 depended on knowing that the DNA structure was a double helix. How can such a revolutionary and fundamental insight of reductive science have had such a delayed impact on our major health problems? Today we are just beginning to sec a flood of practical applications in ‘the pipeline, and one or two have materialized. The molecular genetic prenatal diagnosis of sickle cell disease is one of the first medical appli- cations that explicitly depends on the knowledge of DNA structure: Y. W. Kan’s work is an epitome of the DNA revolution. The biotechnology industries that are founded on recombinant DNA likewise depend on that reductive base. Even with appropriate skepticism about the pace of development of these industries in the next year or two. no one doubts the large number of forthcoming therapeutic innovations. Human proteins such as pituitary hormones, interferon, insulin—and many others today unknown—are accessible in no other fashion. So the texture of my question has changed in the last few years—an authentic turning point in our perspective of history of this phase of medical science. Let me state it 4 bit differently: the phase of application having arrived, why did it take so long? or necd it have taken so long? Some people think such a question is both impatient and petulant, but I - think it ought to be addressed. Over the last thirty or forty years of medical history, one can, of course, trace a host of important innovations. The whole style of medical practice has sharpened, and it is far more attuned to critical scientific inquiry. Physiological and metabolic inquiry, to understand disease process and 203 ACADEMIC MEDICINE management of the care of the patient, from the informed perspectives from immunology and endocrinology as well, is a new common standard. Looking for more specific indicators, I have had some trouble trying to authenticate the most important specific changes in medical practice during that period of time. Once one gets past the antibiotics, which may be regarded as the culmination of the last prereductive era of medical science, it is hard to find a predominant single item in the modernization of medical care. The use of steroids ranks high, despite caveats about latrogenic com- plications. As is typical of many innovations, these complications now loom far larger than first expected. In any event, the initial discovery of the use of steroids in medicine, as with other advances, was closer to serendipity than reductionist planning. My own conjecture is that one of the most important changes in medical practice is the management of the body fluids. I have had some difficulty, however, in getting quantitative data on the history of medical practice with respect to routine fluid infusion therapy. Few will question that this therapy has been a life-saving addition to the armamentarium, if only for the infantile diarrheas. Drinking saline water may in fact become an equally efficacious medical technology! Water does not sound like a very sophisticated medical entity. There are a few things one puts into the water, but they are not particularly complex from a chemical standpoint, and I doubt that one would invoke reductive biology as the route of discovery in this field. But it is all the more reason to seek the different threads that have informed medical practice. We do lack the kind of critical history that would enable us to judge what has happened there, as well as in many -other important changes in practice. Paul Beeson’s comparison of textbooks of medicine is an indispensable way of looking at medical history; but it is almost too comprehensive, and few people will take on the assessment of the most important improvements. In my own view, we are secing, in this decade, the completion of two cycles of medical science and practice. With the DNA revolution we are well into the third. The first cycle rested on the scientific foundations of medical mucrobi- ology laid just a century ago. This was based on the specific recognition of germs as living organisms and as agents of disease: the methods that we owe to Pasteur and Robert Koch, the taxonomy of microorganisms, obtaining them in pure culture and identifying them as etiological agents, the development of vaccine prophylaxis, and antimicrobial therapy, This cycle represented a revolutionary scientific as well as médical finding. It took from 1880 till the 1940s and 1950s to approach an asymptote (Table 1). We well know how mortality from infectious disease has changed since the turn of the century. While, indeed, much of that change can be 204 J, Lederberg CYCLES IN BIOMEDICAL RESEARCH TABLE t Three Major Cycles of Biomedical Progress Cycle Dates Description Develop ments Infectious 1880-1940... Reductive—germ Vaccines disease theory Antibiotics Iluman phys- = 1922-1980... Reductive in atm, Insulin iology ** convergence semi-empirical in Cortisone 1980s practice Diuretics Psychotrop- ics Molecular bi- 1944-1980" Reductive! Enzyme in- ology hibitors DNA diag- nosis Atheroscle- rosis Cancer Transplant rejyecuion attributed to larger cultural and social developments, it is not a question of “either/or”; and one can hardly dispute the importance of scientific knowledge about what. is contaminating our water supplics, or about which vaccines would be effective. Our standards and expectations are much higher today. Even if, after earlier successes, the opportunitics for rapid public health improvement are less today than sixty or eighty years ago, we do not want to stop now. Just think how deprived we would be if we had to rely on these very general measures of sanitation and vaccination, and were barred from the much-derided high technology of medical care. The second cycle I would date to about 1922. It evokes the names of D. D. Van Slyke and J. L. Gamble, i.c., systematic application of human physiology and chemistry in medicine. Many of the specific interventions that are part of medical and surgical practice stem from physiology: the understanding of what the various organs of the body do and how they communicate with one another. Physiology, like much of biology, is informed by medical observations and vice versa. It deserves more honor than it now gets, judging from the departmental arrangements at many of our medical schools. Perhaps just because so much physiology has been incorporated in internal medicine, there is a structural problem fitting physiology as basic science into the organization of many medical schools. 205 discovery has been more significant. An outstanding example is seen in contemporary psychiatric medicine. One cannot describe the development of the now indispensable agents used in the treatment of schizophrenia and depressive illness as having stemmed in any way from a reductive model. Quite the contrary! The empirically demonstrated efficacy of agents like chlorpromazine and lithium then demanded the attention of investigators into the biochemical foundations of the mode of action of the drugs. Their discovery was empirical and preceded the neurochemical theory that is just now emerging. It is a consequence of our Successes against infections that now our The inherent intricacy of these Problems, which are rooted deeply in the molecular and cellular structure of the human organism, outreaches the existing base of applicable scientific knowledge. This ignorance has frustrated the building of a theoretical Program for the control of these diagnostic machines, and by the development of Scientifically trained, sophisticated specialties to make these accessible to patients. This tech- nological revolution has also carried a heavy price tag, and there now experiments, to define the Proper scope of these interventions, to search for their side effects. and so forth. This ramification is, in a way, as indispensable as the initial discovery. It is reaching down: toward the development of a reductive infrastructure for medicine, rather than having built on a deductive foundation for the initial discovery of these useful 206 J. Lederbery CYCLES IN HIOMEDICAL RESEARCH agents. With the exception of prenatal diagnosis, which did start from first principles of genetics and the cytogenetics, very few medical advances have been conceived from prior knowledge of the biology of the organism. My question about the double helix relates to the third cycle, just at its zenith of scientific accomplishment and burgeoning potential for appli- cation. In times past, I might have leaned on the problematical structural relationships of basic sciences to clinical medicine, to account for the imputed delay. The question, it has become apparent, understated the complexity of the task. To illustrate an essential cellular organelle, the ribosome of Escherichia coli (none of this my own work) is sketched in Fig. I. These cartoons show the structure of the ribosome from four quarters. The important point is that the ribosome is composed of no less than 55 different protein subunits. Ribosomes tend to fall apart into a 30 S and 50 S major component. The S’s and L’s are on the two respective columns. At this point, every one of those has now been isolated. The amino acid sequence of the majority has been worked out, at least in some degree. Especially revealing is the self-assembly of this organelle: if you mix the different protein constituents with three molecules of specific ribosomal RNA, the ribosomes will self-assemble from these parts. Whatever magic is in the structural organization of the cell derives from the chemistry of its parts. But what complex chemistry! The extraordinary effort that has been required in order to get to this Stage of knowledge has involved: the mechanical labor of developing methods for the purification of these particles; the separation of their protein constituents in ways that do not chemically alter them; and the analysis of these particles, one by one, in order to determine their chemical composition—always in such a way that their biological integrity would not be degraded. Rather than being impatient about it taking from 1953 until now, one marvels that it has been possible to go that far in the molecular dissection of this very important particle. So it was not enough to proclaim that the structure of DNA was a double helix and to learn the code by which protein structure was determined. That was the revolutionary opening of the door to a vast atray of further investigations of the amazing variety of structures in the cell. From these, one can then expect to see a varicty of applications in human pathology. We alrcady know of genetic diseases of bacteria that result from mutations in different ribosome constituents. Environmental factors also influence ribosomal structure and function. Analogous human diseases are bound to become evident, following the same principles, Unfortunately, there remains a host of technical problems in trying to do the same thing with the ribosomes of eukaryotes. A few of the units have been found. The general structure of the ribosomes is not fundamentally different, but in this case, we must fish these things out of cells that have 207 BAS HED VSS Pema n me ae Ma stn ae CAL ORAM PHENOL GASES 2090-2700 PugOMTtin Peon cr 125 Awa WLMETHYL~ Hag Amo GUAMOSINE THOS EP TOM “7 rains W*-CIM) YL ACENO INE 16S RNA TG RNA UP-G BiROING WHE Vs tear 1 y-twor CUT Domaine 30S SUBUNIT 50S SUBUNIT 30S SUBUNIT 505 SUBUNIT Ribosomal Proteins Pratelns of 30S Ribosomal Protelns of 50S Ribosomal Subunits Subunits Designatlon Mol, Wt Blanding Designation Mol, Wt Binding $1 65,000 Lt 22,000 $2 27,000 L2 28,000 + $3 28,000 U3 23,000 $4 25,000 + 4 28,500 $5 21,000 L5 17,500 $6 17,000 L6 21,000 + $7 26,000 + L7 15,500 S8 16,000 + U8 19,000 S9 17,500 9 $10 17,000 110 21,000 Sit Lit 19,000 $12 17,000 L12 15,500 $13 14,000 Li3 20,000 $14 15,000 Li4 18.500 $15 13,000 + L1S 17,000 $16 13,000 16 22,000 + $17 10,000 liv 15,000 + $38 12,000 L18 17,000 + $19 14,000 L19 17,500 + $20 13,000 + L20 16,000 + $21 - 13,000 L2) 14,000 L22 17,000 Sum 405,000 L23 12,500 + L24 : 14,500 + L25 12,500 + L26¢ 12,500 L27 12,000 (28 35,000 L29 12,000 L30 10,000 U3] Looe L32 133 » 9,000 (34 Sum 549,000 Fic. 1, Ribosomal subunits of Escherichia coli, reproduced with permission of authors and publishers. The four illustrations are from H. G. Wittman. “Architecture of Prokaryotic Ribosomes.” Annual Review of Biochemistry $2(1983):in press. The tabular material is Crom DE. Metzler. Biochemistry. The Chemical Reactions of Living Cells. New York: Academic Press, Inc..1977, n. 929. (4 plus sien in table indicates direct binding to ribosomal RNA.) J. Lederberg CYCLES IN BIOMEDICAL RESEARCH a lot of aggressive enzymes, which tear things apart as soon as they are taken out of their normal niche. I conclude that it is asking too much to expect reductive advances in medical practice until we can fill in the infrastructure between information that is in the DNA, and the way the cell is finally designed and built. Without correctly assembled ribosomes, proper protein synthesis in the cells cannot continue. Ribosome assembly also presents an exciting chal- lenge from the standpoint of its regulatory mechanisms. Here there arc fifty-five different proteins, whose synthesis is precisely coordinated. One finds hardly any unassembled leftover constituents within the E. coli cell under a very wide range of conditions. Some of the protein constituents are able to turn off the synthesis of others at various levels, some at transcription and others at translation, and in that way the system is kept in elegant balance. The details of these interactions again involve intricate geometrical and physical patterning of the reacting macromolecules. Our knowledge of this organelle is matched in some measure by what we know of how cell membranes and several other organelles are put together. However, the cell membrane is not a homogencous, chemically consistent structure, and thus it presents still further challenges to eluci- dating its adaptations to the various roles it must play for different kinds of cells in their own circumstances. Further glimpses into “complexity” come from work on a single bacterium, £. coli. Again, a very important part of the message is that in a comprchensive presentation, the details are unreadable. Figure 2 shows the £. coli genomic map as of two years ago. About 1,000 genetic factors have been identified in E. coli, each known well enough to admit the name of a protein or some enzymic or regulatory function. Most of the morphogenetic variants in the human species would not qualify so we'!, because of ignorance of the protein or regulatory process involved. This map is organized into 100 intervals called “minutes,” in the E. cuit jargon. The reason for such a unit is that the process of fertilization, ic. the transfer of genetic information from a male cell to a female cell, is rather prolonged in £. coli; it takes about 100 minutes for entry, from the beginning of the chromosome to the end. Jacob and Monod showed us how to use the time of entry of a gene for mapping. Finer methods which, in increasing measure, comprise the direct examination of DNA sequences are available today. These hundred minutes of E. coli correspond to about 4 million base pairs: it would take about 1,000 pages of this book to inscribe them onc by one. So far, we know sentences, here and there, adding up to about a dozen pages. We can infer from the local density of the map that the £. coli genome has sufficient information to encode about 5,000 different 209 |. 1 pepo* —— [-————~ pro. B Ipea® 6-4 f———~ at P22 }+—— cam |} —~ or gf (rm B) [>—— rat TH a ho f——— ra wee }+——— leu * (roe) }-——— mn” a rime ° mata" fotA,B” Whe haga 10> torot) 154 18-4 A 9-4 Y i" a zo ounl® let” IwR vert ofl ser pda orca 20- 2t- 2244 23 24-4 26-7 27-4 29-4 aro rpea® avec omoF oss” | oye {peph) compl fond wid gh) tated) hvE p———— adh” {hn Bd so tne A (mun) | (ep) ie P~ — pbea® 30 pula pyr [few aid] {ba (ot E} flaw (cops) fav floK fax Ho) Hot. fey [—— Fr? | flan flaz ner | fat plese |} ——~ fod * tobd* r———— pur p———- red bP——-- ht ® fp urna * (enox) tremC} tear * dodR [—— wpe? -———- ire* P———— awl” hema pth” yn Jeni crn cle Pe i ooh a su0.0 }—— tr TV J [14k ESS gaiv 34 32-4 33-4 344 35- 36-4 374 33-4 40 |_—— pnt }— (teg) {rem} iter} 7 Cotransduchon t———- rol B 4+——- hog” mond bP wae p——— odd tyeS }—— pda +——- pp” 1. ma amet * rot fh aroD f—— —— Foe | btuc — ew phe -——~ int [thes — pas tha paca, 404 + pled Adght torgSt —~ oda Co od ~r0f 4t 4 ew tglywt f——— tor ta-——3t Psi 4 =< | ene fa 42+ 43-4 b-—_~ ie F< tel f1b8 pgs wer fad L—— flan Ne i (gina) lasnT} écm -——— eupd t+—— sup® f———— otk” — Hu? i she onP2H ~~ frerey b———— ew” 44-4 we 45-4 [_—— od non [wan {dcd) SoraF) (matG) A $6 -f-—_ It (oatC,a,0} 47 -]—— 100" 987 woas 494 4 50 ———- ommp* oto ——> [-——— gyrA ubiG gieT apa ——— men B pach * a aE ® pio® ted) —— pwF > --— pd ~ taba -pemB" =~ oC chea moiB- mata a z mR * al 21 4THaIOS 1fbA.8,0 mglABCD Fic. 2. Linkage map of Escherichia coli K-12, Reproduced with permission from Bachmann and Low. Microbiological Reviews 44(198U): 1-56, 50+ Seq 534 5444 5574 aro ———— lod. hat" arlPA-2° —- —~ ded At ded b (nw F) (uxaB) |———~ dave | ~~ dopa }-———— pare ps f——— Pow | — ova—{et 1A [;-——~ hee S }-——— sv” (oath otyA ——— purl ——— onl” rhe pdas® nadh (mC) rana® ung }————- pen feng. | en FENG ev Ad ann [rer ——— pS +—— per TT ORE bates 57—}—+ poocrot 58-4 59 1 aleS ——— roca mam - att + = eabe* ~ mts, {ternOt (pra fl =-- pyG* 7 elk }— — tue 60-- ~~ — tyex — argad 60 61-4 63-4 64-4 65-4 66-4 69-4 4 rot: - argaé r0cB pp 10CC thy fee UM SrreetZ ec) Sy an ee grat * Pa qmed® a give Fa lige} nn an bent re abt . af Aas apes, B }—— anda® L-——— math wip” +-—— spec gic eabe mate ———— ota? (pc B) leit 704 7444 75-4 76-4 739-4 argR ond rt) lo erat” leu alo reO fv J,K ane? on IginT) Colronsducthion gop p———-— high [—~ fou — bis L—— gts” fp aA wy pom fer -~—— f¢na® 80~ 80 ~ 90-4 meta “LL mit eI oce iP }————— pave” ie IA ——— glt€ }———— mret4 4 cy 4 apeA tla® ———— Iya [-——— pal ralG aif 9 | 4—— mai rot ube AC hea tora homes }__. dnoB [dae 4{— a pe } > tyre ee [a gk? 92-4 }___ mgt 4 fotrk metA,8 93-1 Gymat Oewl eri mop ital ——— ono CA jowot fda 04 tft rnc tetd 944 tly [——~ hem" *| ped we i | nn Ws Ot em matt. ER 4 rea reg" Ns oy \\ wer ——— ot S heen * 4 Jobe * | F BST Witt 95 rat Kyte) \\ cor? rpil Ket) \ pid A ye Pb mete vdp* 7 wB,D,E ° 4 tdp foda,e* chlB" ere . fesa* Jour corB 86 tena t 96 by B pola [ra oral b gina, ert val ottP2a" supP (men (rem) 87 97-4 mn ft «