Radioactivity in the Service of Man Rosalvn S. Yalow Solomon ~ A. Berson Research Laboratorv. Veterans Administration Medical Center. Bronx. NY 10468 Department of Clinical Sciences, ~lbert'knsteinCollege of Medicine, Montefinre Hospital and Medical Center. Bronx, NY 10467 ~
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The medical specialty that deals with the application of radioactive materials in clinical diagnosis and therapy has acquired the name "nuclear medicine!' This is a very recent speciality. In 1946,the Manhattan Project was supplanted by the Atomic Energy Commission. As a consequence, radioactive materials (radioisotopes) prepared in the government's nuclear reactor in Oak Ridge became readily and inexpensively available to civilian investigators. This initiated the burgeoning of applications of radioisotopes in medicine. Radioactive materials had previously been employed in medicine. In the 1890's, soon after the discovery of radium by Pierre and Marie Curie (for which they received the Nohel Prize in 1903) radium began to he used in the treatment of cancer. The development of the cyclotron by Ernest Lawrence (Nobel Laureate in 1939), the discovery of the neutron by James Chadwick (Nobel Laureate in 1935),and the discovery of artificial radioactivity hy Frederic Joliot and Irene CurieJoliot (Nobel Laureates in 1935) permitted extension of radioisotope methodology to improve our understanding of human physiology and clinical diagnosis. However, cyclotron-produced radioisotopes were expensive and not readily available. For example, a therapeutic dose of radioactive iodine for the treatment of thyroid cancer cost about $1500 when it was obtained from a cyclotron; an equivalent dose cost $50 when radioiodine was first made available from Oak Ridge. I joined the Veterans Administration Hospital in the Bronx in 1947 to set up a Radioisotope Sewice to make this new modality available to our veteran patients. Thus, my professional life in this field coincides with the lifetime of the field itself. In 1950 Dr. Solomon A. Berson, an internist, joined our Service upon completion of his residency a t our hospital. This began a 22-year partnership that was terminated by his untimely death in 1972. Serendipitous Discovery In common with many other investigators a t this time, our early studies were concerned with the use of radioiodine in the study of thyroid physiology and the diagnosis of thyroid disease ( I ) . I t has been estimated that between 1948-1968 200,000 people with overactive thyroids (hyperthyroidism) received radioiodine, 1311, for the treatment of this disorder; in the same period more than 2 million people received much smaller amounts of radioiodine (tracer doses) for evaluating whether their thyroids were functioning abnormally, Later in this report, I will discuss whether harmful effects of this use of radioisotopes in diagnosis and therapy have been obsewed. We next turned our attention to the measurement of the amount of blood in the circulation. This consists of two components: red cells and a fluid called plasma in which the cells are suspended. This was accomplished by labeling the plasma proteins or red cells with radioactive materials (2). I t should be appreciated that when a sample of blood is taken from your fingertip to determine whether you are anemic, all that is being measured is the hematocrit, i.e., the relative amount of red cells in the circulation, not the absolute volume of either of the two components, plasma or red cells. Measurements of blood volume and of other constituents of the hody are made using what is called the "isotope dilution" method, first de-
BLOOD VOLUME Control~
Anemia
.
Cimhoris
Heart Foilure
Figure 1. Plasma and red b i d cells (RBC) volumes per kliogram of M y mass in control subjects and in patients with anemia, cinhosis, or heart failure. Note that red b l d cell volumes are increased in those with cinhosis or heart failure. The apparent anemia associated with those conditions is amibutable to a more marked expansion of plasma volume.
scribed by George von Hevesy (Nobel Laureate in 1943 for the study of chemical processes). The method requires injection of a radioisotopically labeled substance into the blood and taking a sample of blood a t a subsequent time when the injected material has mixed uniformly in the appropriate hody compartment-about 10 minutes for blood volume determinations. Thus, if the hematocrit is low, it is possible t o determine whether this deficiency is due to an absolute decrease in the number of red cells, i.e., true anemia, or whether it is due to an increase in plasma volume which may occur in several diseases such as cirrhosis or heart failure (Fie. .. 1). . From our work on determinution of plnsma volume, we also noted that the l a l d r d irrurn nmteins I I S P ~in these studies were eventually distributed into a larger body space (3).This led us to develoo mathematical analvses and exoerimental methods for studying the rate of remo;al of album& and other serum oroteins from the blood stream and for determiniw the rates or synthesis and degradation of these proteins (4). f i n s , it appeared reasonable to attemot to aoolv similar methods to &dy the distribution and degradat&in normal and diabetic subjects ot a much smaller protein (or peptidr hormone), insulin. The late I h . I. Arthur Mirsky had suggested ( 5 ) that dialxtesin theadult might becaused hy abnormally rapid degradation of insulin by an ewyrne he called i nau . I'inase. This enzyme is widely distributed in the body. Whv did Dr. Mirsky think diabetes might he associated kith &normal degradation of insulin? It was already well known 30 years ago that, unlike juvenile diabetics whose pancreases contain very small amounts of insulin, the pancreases of the adult-onset diabetics contain almost normal and sometimes even supraVolume 59
Number 9
September 1982
735
DISAPPEARANCE OF I3I1-INSULIN FOLLOWING I.V. ADMINISTRATION
LABELED ANTIGEN
+
Ag'
(F)
LABELED ANTIGENANTIBODY COMPLEX
SPECIFIC
ANTIBOW
Ab
+
A
-
Aq'-Ab (B)
A X b UNLABELED ANTIGENANTIBODY COMPLEX
Figure (RIA).
3.
Competing reactions that form me basis of radioimmunoassay
TIME IN HOURS l d,
Flg~re2 Pepltde-oouna radloanlv ry to plasma as a function of I me fo loumg lnnavenOuI aamtn srrat on of 'l'l-labelea m u m to msulm.weated and Areatw SLOleCIS Tne alsppearance *as retarded in tne lnsul n lrealed rublecls r r o Spective of whether they had receivad the hormone for treatment of diabetes Or forshock therapy for schizophrenia.The retarded rate is a consequence of binding to insulin antibodiesgenerated in response to adminisnation of animal insulins. Note the slower disappearancefram the plasma of MN after 4 months of insulin therapy (curve MNp) than prior to such therapy (curve MN,). (Data reproduced from reference 7.) normal amounts of insulin (6). At that time, it was univenally held that all diabetes was associated with an absolute deficiency of circulating insulin. Thus, it seemed very logical to sueeest that if the pancreas contained sufficient insulin and there was too little-in the circulation, this could he a consequence of abnormally rapid destruction of insulin. T o test the Mirsky hypothesis, we administered radioiodine-labeled insulin intravenously to diabetic and non-diahetic patients. Toour surprise wenbserved just iheopposire of u,hat the Mirskv. hypothesis wuuld have predirted-thrre .. was a slower, not a more rapid, rate of disappearance of labeled insulin from the circulation of nearly every diabetic subject (Fig. 2) (7). Upon closer examination of the history of the subjects, we noted that the differences between those &th a slow &d those with a rapid rate of disappearance was due, not to diabetes per se, but rather to a previo& history of insulin therapy. Included among the "slow disappearers" was a patient who had received massive doses of insulin for shock therapy, a treatment that was commonly nsed in the 1950's for schizophrenia and other mental disorders. Since these studies were performed before the availability of oral hypoglycemic agents (pills which reduce hlood sugar), most diabetics were treated with insulin. Therefore, there were only a few diabetic patients among the MN. who came to us as a fresh "raoid d i s a ~ ~ e a r e r sPatient ." diaietic, wi; a "rapid disappeare;" initially, but converted to a "slow disappearer" following several months of insulin therapy. We, therefore, considered that the reason for the slower disappearance might be the binding of insulin in the blood to antihodies which developed in response to exposure to a foreien substance. the beef-pork insnlins used for treatmerit.
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We appreciated that the failure to have detected circulating insulin antihodies earlier, in spite of the fact that insulin had been used therapeutically for 30 years, was due to the relative insensitivity of classic immunologic methods. Therefore, we proceeded to develop new and highly sensitive methods dependent on the useoiradioiodine-lahi14insulin toquantitate the conrentration of i n s u l i n - l ~ i n dantihodies ~n~ in the rirrulation. We soon appreciated that the same methods used to quantify antibody could be nsed reciprocally to measure antigen, in this case the circulating insulin. Thus, the radioimmunoassay (RIA) principle was formulated as a serendipitous 736
Journal of Chemical Education
1
STANDARD CURVE GP 438
1:100.000 DILUTION OF ANTISERUM
1 2 3 4 5 10 15 pg/mi -2 0.5 1.0 1.5 2.0 2.5 5.0 7.5pM SYNTHETIC 15-LEUCINE GASTRIN I
Figure 4. Standard curve fw the detection of gastrin by RIA. Note that as linle as 0.2pg Of gastrin per milliliter of incubation minure (0.1 picomolar) is readily detectable. (Data reproduced fram Yalow. R. S., Science, 200, 1236 (1978).) fall-out from an unrelated study which asked a question concerning degradation of insulin in diabetics (8). Radioimmunoassay (RIA) is based on a simple principle (Fig. 3). The concentration of an unknown substance is determined by comparing the inhibition of binding of labeled antigen to antibody with the inhibitions observed with a set of known standards. A typical RIA is performed by the simultaneous preparation of a series of tuhes containing standards and unknown samples to which are added fixed amounts of labeled antieen and antihodv. After an a~orooriate time, usually ranging fpom hours to dais, the labeled antigen that is bound to antibody (B) is separated from that which is free (F)and the radioacti\.ity asst8ciated wirh each fraction is measured. The ratiu of H to F for each standard tuhe is then plotted as afnnction of the known concentration (Fig. 4). The unknown concentration is determined hy comparing its ohserved B/F ratio with the standard curve. Thus no radioactive material is iniected into the patient. In some peptide hormone assays as little as 10-14M is measurable in the presence of million-fold higher concentrapermits meations of serum proteins. The sensitivity of RIA surement of insulin in a drop of hlood compared t o the liter of blood required for earlier methods. RIA made possible dynamic studies of the response of peptide hormones to agents
which might stimulate or suppress them. The first discovery made with RIA was that the adult-type diabetic, unlike the juvenile-type, often has a higher than usual level of insulin in the circulation (9). The elevated hlood sugar in these subjects is not caused hy an absolute deficiency of insulin hut rather by a diminished responsiveness to the circulating insulin. A Host of Applications
Among a host of other applications of RIA in studies relating to peptide hormones are: (1)determining whether the small size of certain children is due to their having an inadequate amount of a hormone concerned with growth (growth hormone) in which case their growth rate could he increased by treatment with this relatively scarce hormone obtainable only from the human pituitary a t autopsy; (2) whether excessive steroid production by the adrenal gland is due to a tumor of the gland or to a message from an overactive pituitary; (3) whether or not sterility is due to failure to produce or properly secrete sex hormones; and (4) whether or not a high level of calcium in the blood-which often leads to kidney stones-is due to excessive secretion of the calcium-regulating parathyroid hormone. RIA methodology has since extended beyond the study of the regulation of hormone secretion into virtually all areas of medicine. For instance, it has become the method of choice for screening of hlood in blood-hanks for possible contamination with hepatitis B antigen, the virus which in the past was responsible for 95%of the cases of post-transfusion hepatitis. An application of major importance in public health is the use of RIA in state and regional screening programs t o detect under-activity of the thyroid gland in the newborn. In our country this disease occurs in 1in 4000 births. If i t is not detected and treated shortly after birth, before it is clinically evident, the resulting mental retardation cannot be reversed. RIA of thyroid-related hormones in a few drops of hlood on filter paper permits detection and early simple, inexpensive and effective treatment. Such screening programs would he even more important in other parts of the world, particularly in Southeast Asia, the mountains of South America, or Siberia
where enlarged thyroid glands due to iodine deficiency remains prevalent and the incidence of hypothyroidism of the newborn must be considerably greater. RIA is now in use in thousands of laboratories throughout the world to measure hundreds of substances of biologic interest. A partial listing of such substances is shown in Figure 5-a tahlk which has not heen hrouyht up-to-date since 1975! 'I'he a~vlicationsoi RIA seem limited only . by . the imaaination of the-investigator. RIA uses radioisotopes for in vitro studies. However, much of the specialty of nuclear medicine is concerned with imaging-administering radioisotopes to patients and tracing their behavior in the body. Not only is this methodology an important modality in the diagnosis and management of malignancies, but a new subspecialty, called nuclear cardiology, has achieved considerable prominence. How Low Is Low The henefits of the medical applications of radioactive materials are certainly obvious. Yet the pervasive fear of radiation at almost any level persists. No one can deny that exposure to very high doses of ionizing radiation can kill in days; chronic exposure to more moderate doses also may have fatal conseauences. However. we must also a~vreciate that .. from the d&n 01mankind our progenitors have been exposed to radiation from the natural radioactivitv of soil and food and from extraterrestrial cosmic rays. We have not only survived hut the path of evolution has been uuwards. Thus, exuosure to levelsbf radiation comparable to that of natural backbound has not woven to he iniurious. The remainder of this reuort will address the question as to "How low is low"-or are there levels of lightly ionizing radiation (X-rays, y-rays, beta particles) below which one cannot discern harmful effects? If we want to determine the nature of radiation effects a t dose rates and doses comparable to those attributable to natural background radiation, it would seem most reasonable to examine populations in regions of the world where background radiation is higher than that to which most of the rest of the world is exposed. The usual background dose in most
SUBSTANCES MEASURE0 BY RAOlOIM4UNOASSAY PEPTIML HOREaNES PITUITARY HORMONES G~owthh o m n e adrenocorticotropic h o m n e (ACTHI Helanoyfe rtimulaflng homne ( B H ) .-nSH B-IH
Glyqroteinr Thymid r t l n u l a t i s h o m n c ITSH) F o l l i c l e stimulating h o m n e IFSHI L u t e i n i z i n g h o m n e (LHI Prolacfin Lipotropi" varoprerrin oytocin CHORIONIC HORMONES Human chorionic gonadotropin IHCG) Human chorionlc ~ o n a f m a m o f r o p i n(HCS) PANCREATIC HORMONES ~niulin ~lucagon Pancreatic Polypeptide CALCITROPlC HORMONES Parathyroid h o m n c IPTHI calcitanin l c r ) WSTROINTESTINAL HORWOHES Gastrin secretin Cholcryrtokinin ICCKI varoactive i n t e s t i n a l p l y p e p t l d e (VIP) Gastric i n h i b i t o r y polypeptide IGIPI YASOACTlVL TISSUE HOREaNES Angi~ten~in~ ~radykininr RELEASING AN0 RELEASE INHIBITING FACTORS Thyrotrapin r e l e a r i n g f a c t o r ITRF) LHRF
somatostatin OTHER PEPTIDES Sub5tmce P Endorphins Enkephalinr
NON-PEPTIDAL HORMONES
NON-HOREaNAL SUBSTAIICES
ORUGS k VITAMINS THYROIDAL HORrmNES Cwdix glyco~ide~ Thyroxine I T 4 ) Triiodathyronine IT3) owgr o f ~ b u r e Reverse T3 P l y c h ~ ~ t f vDrugs e STEROIDS Antibiotics AldoIfelOne CNS DCprePIantl vitamin A, F o l i c a c i d c o ~ t i c ~ ~ t ~ ~ o i d ~ CYCLIC NUCLEOTIOES ~~fmgenr Androgens ENZYMES P10ge~feme6 C1 estcmIc PROSTAGLANDINS ~ r w t o s e1. 6 diphorphatare Plaminoge". P l a r n i n BIOLOGIC WINES Chynatryprin, TryPrin serotonin c a ~ b o n l canhydrare irocnrymer neiatonin ~ l d o r ereductare Carboxypeptidase 8 pancreatic e l a r t a r e VIRUSES ~ e p a f i t i ra r r o s i a t e d antigen n ~ r i n c~ e u k m i au l r u r e r ( G m r , Rawher, nolone~l n a r o n - ? f i r e r nankey v i r u s TUEaR UNTIGENS carcinoeabryonic antigen a-htoprotein SERUM PROTEINS Thyroxine binding g l o b u l i n IgG, IgE. IgA. IPM Pmperdin Fibrinogen A p ~ l i p o p r o t e i nB nyoglabin nyelin 8arlc Wotein OMER l n f r i n r i c factor R h e m t a i d factor n a g a n factor Nevrophyrinr staphylococcal B-Enterdtoiln
Figure 5 . Partial listing of peptidal end nonpeptidal hormones and other substances measured by radioimmunoassay.
Volume 59
Number 9
September
of the world is 0.1 rad per year. (A rad is a unit of energy ahsorhed from ionizing radiation per unit mass and corresponds to 0.01 joule per kg.) Such a study was performed in China by examining 150,000 Han peasants with essentially the same genetic hackground and same lifestyle (10). Half of them lived in a region where they received almost threefold higher radiation exposure because of radioactive soil. More than 90% of the progenitors of the more highly exposed group had lived in the same region for more than six generations. The investigation included determination of radiation level by direct dosimetry and evaluation of a number of possible radiation-related health effects including chromosomal aberrations of peripheral lymphocytes, frequencies of hereditary diseases, deformities and malignancies, growth and development of children and status of spontaneous abortions. This study failed to find any discernible difference between the inhabitants of the two areas. The authors of this study concluded that either there may he a practical threshold for radiation effects or that any effect is so small that the cumulative radiation exposure to three times the usual natural background resulted in no measurable harm after six or more successive generations. Let us consider now the regions in the United States where natural background radiation is increased. In Colorado and Utah, the average radiation exposure is about twice that on the East and West coasts because of increased cosmic rays a t their higher elevations and natural radioactivity of the soil. However, death rates due to cancer in these states are among the lowest in the country. I t is possible that an appropriate statistical analysis would reveal that the racial, ethnic, age distribution or other factors might account for the lower cancer death rates in these two states-after all, Mormons neither smoke nor drink. When Mason and Miller (11) compared the age-adjusted risk ratio for mortality from malignancies for Caucasians in Denver and Salt Lake City with those in San Francisco and the county in Washington in which Seattle is located, they ohsenred that the leukemia incidence was slightly hut not significantly lower and the incidence of other cancers was significantly lower in the higher radiation exposure cities. An inverse relationship between elevation (h&e higher radiation exposure) and mortality from leukemias and lvm~homashas also been rewrred (121. Others have concluded(l$) that in the United states there is no relation between increased background radiation and leukemia. There are regions of the world-in Kerala, India and in Brazilwhere, because of natural radioactivity in the soil, the DODUlation exposure is even higher. studies of the population in these regions have also failed to demunstrate radiation-induced deleterious effects (14-18). I t is evident from data such as these that radiation exposures at levels as much as three times and perhaps as much as ten times the usual natural background does not result in any measurable increase in deleterious health effects. Effects from man-made radiation at comparable dose-rates and cumulative doses are expected to he similar to those from natural radia-
"......
tin"
Literature Cited 11) Bemon, S. A., Yslow, R. S., Sonentino, J., and Roswit, 8..J. Clin, lnuesl.. 31, 141 11952). 12) Berson, S. A,, and Yalon. R. S..J Clin Inwsl., 31.572 11952). 13) Berson, S. A., Yslov, R. S., Sehniber, S. A., and Post, J.. J. Clin Inueaf., 32.746 ilPX1, ~.""",. 14) Berson. S. A.. and Yalow. R. S.. J. Clin. Inuesf.. 33.377 119541 ..."~"..,"",.,"\A""",.
161 Yalow. R. S.. and Bemn.. A. A...Nolurs.,~ 1S1.lM.R ~.. (1969! 9, \'*I 1v.H S , s n d n . n ~ m S A . J l'lm 1nlr.r .J9. 11571196111 Ilrt H ~ z h B a . k ~ n w n d Had#m,n R l r l r r h G r r w ~ , V h . n a .'Ilvnlrh S u m y ~n Htyh Bal k. ~
What about radiation effects at slightly higher dose mtea and doses? I referred earlier to the use of ""I for the diagnosis of thyroid disease and treatment of hyperthyroidism. Those treated with l3II were exposed rather acutely to generalized hody radiation significantly greater than that frGm natural background, A study of 36,000 hyperthyroid patients from 26 medical centers, of whom 22,000 were treated with l3II and most of the rest with surgery, revealed no difference in the incidence of leukemia between the two groups 119).The average hone-marrow blood dose due to the 1311was about 8-10 rads, half of which was delivered within one week; the thyroidal dose generally exceeded 10,000 rads. The follow-up for the '3II-treated group averaged 7 years, quite comparable to
738
the delay time of peaking of leukemia incidence (5 to 9 years) following atom bomb exposure. This group of '311-treated patients had no increase in leukemia rate comnared to hvperthyroid patients treated with surgery. This study then;s consistent with the inabilitv to demonstrate induction of malignancy due to an acute radiation dose of about 5-10 rads. Studies such as these involving tens of thousands of people which fail to demonstrate a radiation-induced effect are considered not newsworthy and receive no puhlicity. On the other hand, there are some studies in which a clustering of leukemia or other deaths among small groups occurs simply on a random basis unrelated to any other factor. These studies purporting to show radiation effects a t low doses and dose rates are deemed newsworthy, receive considerable publicity and the public accepts their validity. At present, there is no firm evidence excluding or establishing that there is a threshold below which there are no radiation effects. T o obtain statistirallv valid evidence for such e i i e ~ t would s requireexamination ot tens or hundreds of million . nwple. . This would obviously he an impossible task. With what we have learned in recent years through studies in molrculnr hiulom, there is every reason 10 helie\r thar our bodies have rrpnir mechanisms hitherto undrcamrd of. The disagreements regarding extrapolation of high dose data to the low-level radiation concern hypothesis, not obseruable facts. I might remind the readers that even Newton's Laws, which were considered to be universally valid for more than two centuries, were not appropriate descriptions of nature when extrapolated to subatomic dimensions or velocities approaching that of lieht. While there was no need to make S i c y decisions basedon extrapolation from Newton's Laws, public ~ o l i c vdecisions do need to he made in settine radiation protection standards even in the absence of scienti6 evidence. Under those circumstances a risk-benefit analysis seems the best way to reach such decisions. It should be appreciated that regulatory decisions are often arbitrary, based on philosophy not fact, and may well change because of political or other considerations. There is a distinction between hypotheses and observable scientific facts. Those concerned with potential risks associated with low-level radiation should make that distinctiun. The benet'its of rndioactivitv in the service of man are real and significant. Let us not he kept from these benefits by irrational fears generated by well-intentioned or ill-intentioned hut often uninformed Cassandras.
Journal of Chemical Education
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