ML13220A065

LTR-13-0660 - Edward Calabrese, University of Massachusetts Email Two Papers Entitled Origin of the Linearity No Threshold Dose-Response Concept and How the Us National Academy of Sciences Misled the World Community on Cancer Risk Assessmen

ML13220A065
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From:	Calabrese E J Univ of Massachusetts - Amherst
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Joosten, Sandy From: Sent: To:

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Dear Chairperson:

Edward <edwardc@schoolph.umass.edu>

Wednesday, August 07, 2013 10:41 AM CHAIRMAN Resource new papers Archiv Toxicol Origin LNT-l.pdf; Arch. Tox. NAS-l.pdf I am attaching two papers; one is on the origin of the LNT and the second deals with the documentation of how leading radiation geneticists deceived the scientific community on LNT, leading to the US National Academy of Sciences' (NAS) recommendation to adopt the LNT model for risk assessment.

These assessments may be of some interest to you. Sincerely, Ed Calabrese 1

Arch Toxicol DOl 10.1007/s00204-013-1104-7 REVIEW ARTICLE Origin of the linearity no threshold (LNT) dose-response concept Edward J. Calabrese Received:

15 March 2013 I Accepted:

II July 2013 ©Springer-Verlag Berlin Heidelberg 2013 Abstract This paper identifies the origin of the ity at low-dose concept [i.e., linear no threshold (LNT)] for ionizing radiation-induced mutation.

After the covery of X-ray-induced mutations, Olson and Lewis (Nature 121(3052):673-674, 192H) proposed that cosmic/ terrestrial radiation-induced mutations provide the cipal mechanism for the induction of heritable traits, viding the driving force for evolution.

For this concept to be general, a LNT dose relationship was assumed, with genetic damage proportional to the energy absorbed. sequent studies suggested a linear dose response for ing radiation-induced mutations (Hanson and Heys in Am Nat 63(686):201-213, !929; Oliver in Science 71:44-46, l ()30), supporting the evolutionary hypothesis.

Based on an evaluation of spontaneous and ionizing radiation-induced mutation with Drosophila, Muller argued that background radiation had a negligible impact on spontaneous tion, discrediting the ionizing radiation-based ary hypothesis.

Nonetheless, an expanded set of mutation dose-response observations provided a basis for ration between theoretical physicists (Max Delbruck and Gunter Zimmer) and the radiation geneticist Nicolai Timofeeff-Ressovsky.

They developed interrelated physical science-based genetics perspectives including a biophysical model of the gene, a radiation-induced gene mutation target theory and the single-hit hypothesis of radiation-induced mutation, which, when integrated, provided the cal mechanism and mathematical basis for the LNT model. The LNT concept became accepted by radiation geneticists E. J. Calabrese (C8:1) Department of Public Health, Environmental Health Sciences, University ofMassachw;etts, Morrill I, N344, Amherst, MA 01003, USA e-mail: edwardc@schoolph.umass.edu Published online: 26 July 2013 and recommended by national/international advisory mittees for risk assessment of ionizing radiation-induced mutational damage/cancer from the mid-1950s to the sent. The LNT concept was later generalized to chemical carcinogen risk assessment and used by public health and regulatory agencies worldwide.

Keywords Ionizing radiation

• Linearity

• Dose response

• Risk assessment

• Threshold dose response

• Target theory

• Eugenics

• LNT Introduction In 1956, the US National Academy of Sciences (NAS) Committee on Biological Effects of Atomic Radiation (BEAR I)/Genetics Panel issued the most far reaching recommendation in the history of risk assessment that genomic risks associated with exposure to ionizing ation should be evaluated with a linear dose-response model, no longer via the threshold dose-response model that had long been the "gold" standard for medicine and physiology (Calabrese 2005, 2009a, 20 l l ). The Genetics Panel members believed that there was no safe exposure to ionizing radiation for reproductive cells with the tion risk being increased even with a single ionization (Hamblin .2007). The LNT concept was generalized in 1958 to somatic cells and cancer risk assessment by the National Committee for Radiation Protection and urement (NCRPM) (Whittemore 1986). Quickly ter, other national and international advisory committees and organizations adopted such judgments for ionizing radiation (Calabrese 2009b ). In 1977, the Safe Drinking Water Committee (SDWC) of the US NAS extended the linear dose-response risk assessment model of the BEAR/

Biological Effects of Ionizing Radiation (BEIR) tees to chemical carcinogens, a recommendation that was soon adopted and implemented by the Environmental tection Agency (EPA). On a parallel track, similar LNT risk assessment procedures were adopted by the Food and Drug Administration (FDA) in 1977 concerning animal carcinogen drug residues.

Despite the fact that the LNT model has been of central importance in chemical and ionizing radiation regulatory risk assessment, its origin is not within the environmentaV occupational risk assessment domain. The current paper provides a novel historical assessment of the scientific gin of the LNT. It will show that the LNT was first applied to the field of biology in 1928 to explain the occurrence of genetic variation that would serve as the "biological engine" for evolution.

The paper will also demonstrate how the linear dose-response model as proposed by Olson and Lewis ( 1928), which soon afterward became transformed into a "Proportionality Rule" by Muller ( 1930), became mechanistically framed within the context of a single-"hit" hypothesis based on the target theory by vsky et a!. ( 1935) in a unique collaborative effort between leading theoretical physicists and radiation genetics.

This paper extends two earlier publications within Archives of Toxicology concerning historical foundations of the LNT concept (Calabrese, 2009b) and threshold/hormetic brese 2009a) models. Evolution and LNT Since the publication of the Origin of Species in 1859 by Darwin and the rediscovery of the works of Mendel on gene inheritance, there was intense interest in the cal community to determine the cause of genetic change or novelty that would be subject to natural selection, thereby providing an important mechanism of evolution.

As noted by Patterson ( 1933). a well-known colleague of Hermann J. Muller at the University of Texas/Austin, "the important question in biology is the problem of evolution" referring to the need to understand the mechanism of evolution at the gene level. Despite the fact that the gene was more of a concept than a physical entity during the early decades of the twentieth century, it was widely believed that the gene was the basic unit of heredity and that the driving force for evolutionary change must be via the induction of able genetic changes or mutations at the gene level ler l 922). This perspective provided the basis for intense interest by numerous genetics researchers in the second and third decades of the twentieth century to induce tions in heritable traits by environmental (e.g., temperature) alterations, physiological stressors (e.g., starvation), as well as toxic chemicals and ionizing and non-ionizing radiation.

Arch Toxicol Given the central importance of evolution in biology and underscoring the intensity of the competition to be the first to demonstrate inducible heritable changes, Muller ( 1927) provided only an initial "discussion" of his genicity findings with no data in his now famous Science paper that led to his Nobel Prize in 1946. This was done in order to secure recognition of being the first to report induction of heritable mutations by an environmental agent (i.e., X-rays). The supporting data were published the next year in a conference proceeding of very limited distribution based on the World Cat database (Muller 192Xa) and also within the Proceedings of the National Academy of ences (PNAS) (Muller 1928b ). Not only were the findings of mutation significant so too was the fact that the mutation rate was increased by about 150-fold at the highest dose tested. Muller speculated that naturally occurring ionizing ation might be a significant explanatory factor for genetic variation and may drive the evolution process. However, Muller was cautious in making the mutation-evolution link as the doses he had used to induce mutation were extremely high, exceeding background by about 200,000-fold, ing sterility or mortality in a substantial proportion of the fruit flies tested. In addition, the dose response was not linear but closer to a square root function due to a est decline from linearity at the highest dose (Muller 1927, 1928a). If the true dose response for ionizing induced gene mutation was linear at low dose, as a general condition, then it may have explanatory implications for an evolution mechanism.

Consequently, he soon directed several members in his laboratory to assess the topic of dose response more fully than he did in his ing mutation discovery.

While the follow-up research by Muller's group was being undertaken, Axel R. Olson and the prestigious physical chemist Gilbert N. Lewis (1928) of the University of California/Berkeley published a posal on April 28, 1928, in Nature that natural ity was likely a significant cause of mutation that could generate variability from the parent generation and affect the process of evolution.

These authors based this sition on a report of January 1, 1928, in PNAS by speed and Olson on X-ray-induced heritable changes in tobacco. These authors claimed that the tobacco plant ies were specially planned to facilitate a direct comparison of mutation rates between the artificial X-rays and rally occurring radiations." Olson and Lewis ( 1928) also stated that "since the rays can only be effective when they are absorbed, and this produces ionizations, it seems safe to assume that the various rays will produce biological effects in proportion to the ionization which they cause" (emphasis added), a perspective based on the emerging target theory for radiation-induced biological effects proposed by leaders in the physics community (Glocker !927; Crowther 1924).

Arch Toxicol Olson and Lewis ( 1928) then utilized a simple linear ematical model to derive a mutation estimate at a selected natural background radiation dose. With this method, they estimated the number of variants (mutants) induced per year by natural radiation.

These authors concluded that "it seems, therefore not altogether extravagant to assume that such variations as actually occur in nature are due largely to the radioactivity of the environment." The ment of Gilbert Lewis in this activity, while unexpected, was derived from his research in the 1920s in the area of radiation physics (Coffey 2008). Furthermore, his eclectic research activities had also drawn him toward ary theory, the subject of his major presentation (i.e., man Lecture) at Yale, just preceding the development of the LNT paper in Nature (Lewis 1926). This lecture followed that of Thomas Hunt Morgan of Columbia University in 1925, Muller's Ph. D. advisor and 1936 Nobel Prize ent. The perspective of Olson and Lewis ( 192r\) was also independently advanced by Muller in a paper read before the National Academy of Sciences on April 24, 1928, and published on September 14, 1928. The statement of ler ( 1928b) was principally conceptual, lacking the detailed formulation of Olson and Lewis (I 928). The following year, Babcock and Collins ( l929a, b) tested the hypothesis of Olson and Lewis ( 1928). They found a location in which the natural radiation was twice that found in their University of California/Berkeley ratory. Using the CIB strain sex-linked recessive ophila assay, they reported an increase in mutation that corresponded in the same proportion as the difference in background radiation, supporting the proportionality hypothesis.

Detailed experimental methods including the actual radioactivity levels were never published, although such data were promised to be provided in a subsequent paper. In 1930, Hanson and Heys provided further support for the hypothesis that "natural radiation may be ble for the mutations that are the grist of the natural tion mill with the resulting evolution of new forms." Their findings were based on a study of fruit fly mutations in an abandoned carnotite (i.e., uranium) mine. Such tions were initially supported by commentaries by various authors (Lind 1929; Dixon 1929, In 1930 Muller and Rice University physicist, Smith, challenged this LNT evolution perspective by reporting that natural radiation, which was of such a dose rate, could only account for about Ill ,300 of the gene mutations that occurred spontaneously in Drosophila nogaster, assuming a linear dose response.

The authors concluded that other causes must explain the origin of most mutations that spontaneously occur. Nonetheless, in his sertation, under the direction of Muller, Oliver ( 1931) stated that cosmic and terrestrial radiations must account for some proportion of the spontaneous mutations (see Muller I 930). This conclusion was justified on the belief that the response is linear at low dose, with there being no threshold for a mutation response.

This relationship was stated as holding true for all types of high-energy radiation (e.g., gamma, beta, X-rays and probably ultra-violet rays). Thus, Oliver ( 1931) concluded that "by inference it can be added that the cosmic and the terrestrial radiations also are capable of producing mutations in proportion to their power of tion." Oliver ( 1931) also extended the concept of tionality to chromosomal inversions and translocations further arguing for the support of a background radiation influence.

For example, Muller and Altenburg ( 1930) noted that translocations are induced at a similar frequency as gene mutations.

Given these circumstances, Oliver ( 1931) noted that "one would expect each of the classes of changes considered to occur with the same frequency when the viduals are subjected only to the natural conditions, if ral radiation can account for all mutations

... " Despite this interpretation of environmental radiation-induced genetic changes, Oliver ( 19 31) concluded that "some other tion must, therefore, enter in order to explain the difference in non-radiated material, between the frequency of gene mutation and that of the other type of genetic changes." (p. 34) Even though Muller dismissed natural radiation as viding a quantifiably significant mutational influence to derive genetic novelty for evolutionary change, he still retained his belief in the linear dose-response relationship (p. 238) (Muller 1 930) based on the findings of Hanson and Heys ( 1929, I 930) and Oliver ( 1930). Even though the hypothesis of Olson and Lewis ( 1928) did not maintain nificant support for long within the scientific community, Muller and other leaders of the radiation genetics nity became strong advocates of the LNT model to account for genomic mutations and the occurrence of cancer. It may seem difficult to understand in retrospect why prominent scientific leaders such as Gilbert N. Lewis, mann J. Muller and others so quickly adopted a belief in linearity at low dose. In the case of Muller, he was fully committed to this view after the publication of only three studies (Hanson and Heys 1929, Oliver 19.i0) in which the lowest cumulative dose was roughly 285 r, administered in an acute manner, the rough tion of 1,000 modem chest X-rays in 3.5 min or 5 chest X-rays/s.

In his rather copious publications during this period of "belief'/concept formulation, Muller never addressed temporary publications that did not support a linear pretation (Patterson 1928; Weinstein

! 92X; Stadler I 9:10, I tn I). Yet, he was well aware that the lowest doses in the Hanson and Heys ( 1929, 1930) and Oliver ( 1930) papers were acute studies that grossly exceeded background tion exposure.

To think within a linear dose-response term framework ran counter to pharmacological and chemical toxicological experience at that time. As Zimmer ( 1966) reflectively wrote, toxic chemicals in the early decades of the twentieth century demonstrated "no effect up to a threshold dose and then climbed steeply up to 100 %." Muller and others argued that the genetic response to izing radiation demanded a different evaluative framework.

Target theory and LNT A likely explanation for Muller's (and possibly Gilbert N. Lewis's) acceptance of the LNT in the absence of ing dose-response data may be found within the scientific culture at the time. X-ray-induced mutational effects were placed within the context of what was called the tion target theory. This theory was quantitative and metric, with mathematical calculations related to quantum mechanics, reflecting the leadership of prestigious retical physicists (von Schwerin 2010). The formation of a physics-based target theory was established prior to the covery of inducible mutations by Muller ( 1927) by cal physicists such as Dessauer ( 1922), Glocker ( J 927) and Crowther (!924, [926, 1927), setting the stage for a novel scientific framing of the mutational data in the 1930s. The mutation findings of Muller ( 1927) were a major scientific advance that easily fit into the target theory concept while also markedly advancing the scientific standing of target theory itself. The radiation target theory as applied to mutations was formulated by the detailed interactions and collaborations of leading radiation geneticists and theoretical physicists during the mid-1930s.

During this time, radiation cists, lead by Nicolai Timofeeff-Ressovsky, and cists, including Niels Bohr, with a profound interest in the interface of physics and biology, would meet each year, typically in Copenhagen and Belgium for extensive sions. From these exchanges developed the seminal ceptual paper by Timofeeff-Ressovsky and the physicists Max Delbruck and Kevin Gunter Zimmer ovsky et al. 1935) that would establish a conceptual work for gene structure, target theory for the induction of mutations via ionizing radiation, the single-hit nism hypothesis to account for the shape of the LNT dose response and the application of this dose-response model for what was to become modern cancer risk assessment.

The genetic target theory saw mutation as a purely physical action following an all or none Jaw in which a single zation or energy absorption produces the mutational effect independent of all other ionizations and energy absorptions.

This linearity feature stands in contrast to normal ology that invariably deals with large numbers of ecules of each kind, and where the elimination of a single 'f) Springer Arch Toxicol molecule would not result in observable effects (Delbruck J 940). The energy of ionizing radiation was assumed to be essentially transformed into a genetic effect. According to the physicist turned biologist Max Delbruck (1969 Nobel Prize recipient in Biology and Medicine), the ality rule that was proposed earlier by Muller, based on the research of Hansen and Heys ( 1929) and Oliver ( 1930, 19 3 1) and supported in experimental research by eff-Ressovsky et al. ( l 935), provided the basis of the gle-hit mechanism interpretation and the calculation of the size of the gene (Delbruck l9..J.O).

Table I provides a ing of quotes in which the early conceptual framing of the dose-response proportionality concept occurred.

The forming of a dose-response hypothesis based on a very limited amount of data into a biological "Rule" by Muller was done without significant discussion of the concept, its possible mechanisms as well as the recognition of data that may contradict this "Rule." Although Muller was a geneticist, he was drawn quickly toward the physics-mutation interface, accepting significant elements of target theory for radiation-induced mutational effects, including the important assumptions that age was proportional to the energy absorbed, linear response modeling and that effects were cumulative and deleterious (Muller et al. 1936). Muller knew Ressovsky, having met him in the Soviet Union in 1922, encouraging him and his colleagues to transform his ratory to one of the Drosophila genetics.

Muller renewed contact with Timofeeff-Ressovsky during the 5th national Congress on Genetics in 1927. From November 1932 to September 1933, Muller researched in Berlin with Timofeeff-Ressovsky.

He also participated in the biology/mutation discussions in Copenhagen in 1936, engaging Niels Bohr and other leading physicists. ments of radiation geneticists during this period were often designed within the context of this target theory framework.

This was also the case for critical studies performed a ade later under the aegis of the Manhattan Project at the University of Rochester under the direction of Curt Stern (with Muller serving as a consultant) (Spencer and Stern 1948; Caspari and Stern 1948). The hit hypothesis As noted above, in his Nobel Prize research, Muller reported that the induction of mutations was not directly proportional to the X-ray dose, but rather to the square root of the dose (Muller J 927). Based on discussion with the physicist and future Nobel Prize winner Irving Langmuir (1932 Nobel Prize in Chemistry), Muller ( 1927) stated that this observation suggested that the induction of tion was not caused directly by a single quantum of energy.

Arch Toxicol Table 1 Documentation of the introduction of the proportionality rule concept into the mutation literature, 1929-1960 References Hanson and Heys (I '129) Muller ( 1930) Oliver ( 19 30) Stadler (

Hanson et al. ( 1931) Oliver (1931) Oliver(l93l)

Oliver (I ,I) Patterson ( Fn I) Hanson and Heys (I ()C\2) Hanson and Heys (I <JC\2) Hanson ( 19.'\.<) Oliver (!()34) Delbruck ( 19+0) Stern ( 1950) Stern ( 1960) Quote "It is only to be expected that the number of mutations be directly proportional to the number of rays to which the organisms are exposed." Page 207 "Since then Hanson, using radium, and Oliver in our laboratories using X-rays; have both found that the quency of mutations produced is exactly proportional to the energy of the dosage absorbed ... There is, then, no trace of a critical or threshold dosage beneath which the treatment is too dilute to work." Page 236 "That is there is a direct proportionality between the percent of lethals and the length of time of treatment may be seen more readily by a comparison of the t 1 values calculated from the results for each of the given doses." Page 45 "Mutation frequency increased approximately in direct proportion to dosage." Page 13 "Taking the amount of ionization in air as a measure, the mutation rate seems to vary approximately in direct proportion to the intensity." Page 142 "By inference it can be added that the cosmic and the terrestrial radiations of higher energy content also are capable of producing mutations in proportion to their power of ionization." Page 480 "The relation of proportionality to the dosage applies not merely to the lethals in general, but, more specifically, to the lethal gene mutations." Page 485 " ... [gene mutations and gene rearrangements]

all probably occur in direct proportion to the dosage, no matter how small a dose is used." Page 486 "In general their results [i.e., Hanson and Heys I <J2R and Oliver 19:10] justify the conclusion that the rate is directly proportional to the dosage employed." Page 133 "Further evidence of the proportionality rule from a study of the effects of equivalent doses differently applied." Page 335 "Experiments planned with a view to determining within what limits the proportionality rule holds show again a strict correspondence existing between the amount of radium administered and the consequent biological effect, the induced mutation frequency obtained varying directly with the dosage." Page 343 "The rate seems to be directly proportional to the dosage. Muller has named this the 'proportionality rule.' For example, when all other factors are kept constant, doubling the time of exposure also doubles the number of lethal mutations." Page 486 "The frequency of induced mutations is directly proportional to the intensity of the treatment" Page 391 "The proportionality rule gave the basis for the single-hit interpretation

... " Page 359 "The proportionality rule has been proven to hold over a wide range. Figure 155 shows that, for Drosophila, the relation is essentially linear over the range from 25 r to several thousand r. It has further been shown that the frequency of induced mutations is independent of the time over which the radiation is applied." Page 433 "It has been established for a variety of experimental organisms that the number of mutations induced by tion is proportional to the dose. This proportionality has been proven to hold over a wide range of dosages." Page491 However, subsequent exposure experiments by Hanson and Heys ( 1929), Oliver ( !930, 193 I) and later by Ressovsky et a!. ( 1935), even though all experiments were at very high dose, supported a proportionality relationship, which was consistent with the "hit" theory of mutation in which the X-ray treatment excites an electron in the target gene. This excitation was proposed to affect a permanent change or mutation to a different molecular structure. izing irradiation was the only effective way to induce tions; it showed no threshold, suggesting that the tion of radiation is a quantized and additive process (von Schwerin 20 l 0). A "quantum-jump" was considered to be the physical process caused by a hit on a target, resulting in mutation.

Treatment effects induced by a physical agent like ionizing radiation were believed to be caused by one or several discrete biophysical events, that is, hits on a target. Based on hypotheses about what constituted a hit, cal models were used to construct dose-response ships. If there was only a single hit on a single target, the dose response was linear. As the number of assumed hits increased, a more threshold like the dose response would appear. In a practical sense, the mathematical derived dose response based on an assumed number of hits could be visually matched against the laboratory-obtained dose-response curve. Using this direct and simplified approach, researchers like Muller, Timofeeff-Ressovsky and participating physicists decided the theoretical number of hits. This type of target theory was especially strong in Germany, with support from leaders such as Boris sky (Director of the KWI for biophysics, 1936),

Ressovsky and others (von Schwerin 20 I 0). This tual framework led to the conclusion that mutation was a single-hit process, proceeding from a single ionization, from a quantum of ionizing radiation in a specific sensitive zone of the gene. This theoretically based perspective became not only a workable model but a firm belief within the radiation genetics community even though there was no edge of the physical nature of the gene. As coauthor of the Timofeeff-Ressovsky et a!. ( !9 35) paper, Delbruck quently noted in his Nobel Prize lecture that it was thought that genes were very stable and, therefore, showed teristics of molecules.

However, the gene concept at that time was simply that of Mendelian algebraic rates, lacking structural chemistry insight. There was much speculation of gene structure including that of submicroscopic state systems or even an entity not readily analyzable in chemistry as proposed by Bohr ( !933 ). The paper of Timofeeff-Ressovsky et a!. (1935), as noted above, was striking in its collaboration between physics and genetics, its proposed chemical nature of the gene, size of the gene and in the proposal of a "hit" esis as the foundation of the linear dose response for izing radiation-induced mutation.

While the gene structure and size framework would be bypassed and replaced by the DNA structure of Watson and Crick ( 1953), the hit ory component of Timofeeff-Ressovsky et a!. (! 935) was accepted and implemented by the radiation genetics munity. The term "hit hypothesis" became commonly used in the lexicon of radiation genetics, including those prising the BEAR I Committee/Genetics Panel that mended changing to a linear model from a threshold model for assessing mutation risks from ionizing radiation brese 2013). The impact of this 1935 article was facilitated by the actions of Timofeeff-Ressovsky who sent reprints to key researchers.

However, the overall immediate impact of the paper was very limited as it was published in an obscure Gottingen journal that was not cited in any leading index with only four issues being printed before ceasing tion. This paper, which provides the origin of the single-hit hypothesis to support a linear dose-response model, was not even cited in the BEAR I report that implemented the concept. Yet, the term "hit" hypothesis and target theory became commonly used, even if credit was not often given to the original paper (Timofeeff-Ressovsky et a!. l 93:'i). Nonetheless, this paper did receive a major endorsement in the 1944 book "What is Life" by Erwin Schrodinger, a Nobel Prize physicist (1933), raising its visibility in the physics community.

The concept of the gene and its striking stability gested it must have a unique atomic composition. bruck (I 970) believed that such stability might be due to each atom of a gene being fixed in its mean position and electron-stable, sunk in an energy well, now seen having Arch Toxicol stability due to the function of the hydrogen bond. tions of such genes could only occur following the tion of high energies as from ionizing radiation, not from heat under physiological conditions.

In fact, a modest increase in vibrational energy was estimated to increase the atomic stability, decreasing mutational risk. Since a transaction in an atom can be affected by a single digit e V and that the initial impact of an X-ray can be several fold greater, it was believed that any gene would be at risk for mutation from radiation.

Since the initial energy of impact exceeds a threshold energy of activation, ionizing-radiation should affect not only the induction of a localized mutation but also that of a broad range of gene targets. The mutation hit theory was challenged by Caspari and Stem ( 19-+R) in a chronic, very low-dose rate study, leading to the hypothesis that either a threshold exists or multiple independent primary actions are required for a mutation to occur, or that a recovery or* repair cess occurred at a very low-dose rate (Howarth et a!. 1950; Key 1951). Over the next several decades, the dominance of the physics-based target theory would yield to improved chemical/biological/physiological understandings of the mutation process, including such modified target theory effects of ionizing radiation as DNA repair (in tive and somatic cells), adaptive response, the bystander effect as well as the recognition that the biological effects of ionizing radiation are principally due to the tion of hydroxyl radicals/hydrated electrons from lar water and their migration to cellular targets (Collinson eta!. !962; Czapski and Schwartz !962; Weiss 19-1-4). In fact, even as the target theory was being applied to tion by Timofeeff-Ressovsky et a!. (!935), the recognition of repair processes, including DNA repair, were emerging (Hanawalt 199-1-). Such challenges to the hit theory would eventually be brought to the BEAR Committee by Russell ( J 956, 1963) from Oak Ridge, but only after the BEAR I Committee made its linearity recommendation.

Edward Lewis (J 957a), another radiation geneticist Nobel Prize (1995) recipient, published a very influential Science article in 1957, strongly supporting a linear tionship for cancer, relying on linearity data in the Uphoff and Stem ( 1 949) paper. In subsequent Congressional timony, Lewis (I 957h) would argue that the dose response was linear, regardless of the mechanism, and should be accepted as such whether or not a mechanism could even be discerned.

These comments of Lewis suggested that he ognized the growing mechanistic challenge to the hit theory as well as new conceptual problems (e.g., tiple biological processes could yield a linear relationship that did not require a single-hit process) emerging from the physics and genetics communities, including Zimmer ( 1941 ), a coauthor of the Timofeeff-Ressovsky eta!. ( 19 35) paper and radiation biologists/geneticists (Haas et a!. 1950; Arch Toxicol Kimball I 952). However, the time period within which Muller's mutation were produced was one of the cultural scientific dominance of physics. Association with the leadership of the physics community served to enhance the significance of the mutational findings and its assumed linearity at low dose, as well as providing Muller with an expanded scientific and cultural context that recognized his achievements and enhanced his scientific reputation.

The influence of the hit concept of Timofeeff-Ressovsky et al. (1 935) was facilitated via subsequent publications of Lea ( 1940, 1946), which offered further justification for the target theory-based LNT-single-hit hypothesis for tion. The publications of Lea were not only authoritative extensions of Timofeeff-Ressovsky et a!. ( 1935) but more readily available than the Timofeef-Ressovsky et al. ( 1935) paper with its publication in a defunct journal. Regulatory agency actions Ionizing radiation In the radiation risk assessment area, two endpoints were adopted to which linearity was applied: germ cell tions and cancer. In the case of germ cell mutations, based on several publications in the early I950s by Muller ( 1951, 1954), the BEAR I Genetics Panel (1956) proposed to limit exposure to ionizing radiation such that exposure would not exceed doubling of background mutations from tion through the first 30 years of life. The panel assumed that exposure to ionizing radiation could cause mutations to germ cells in a linear manner and had the potential to cause adverse genetic effects in individuals and future erations.

The panel derived a risk assessment ogy for application to both first-generation offspring and total genetic risk, including future generations.

The panel derived a doubling dose method (i.e., the dose of ionizing radiation, assuming linearity at low dose, that would equal the number of mutations resulting from background sure), to estimate population-based risks. This doubling dose methodology would predict the number of genetic diseases based on three parameters:

the assumed doubling dose, the proposed exposure limit and the background dence of genetic disease. Based on this risk assessment framework, the panel recommended a "uniform national standard" such that the members of the general tion would not receive more than a cumulative dose of I OR from conception through 30 years. This basic method of the BEAR I Committee, using the doubling dose/linear work, has been refined with recent advances allowing one to integrate between rates of radiation-induced mutation based on mouse studies and the risk of inducible genetic disease in people [Sankaranarayanan and Chakraborty 2000a, b; Sankaranarayanan and Wassom 2008 (see Lyon 2003 for an alternative view)]. In the case of somatic effects, cancer risks were mated via the use of a linear dose-response model. ing linearity to zero, it was estimated that exposure of one rem to one million people each year would cause one to two new cases of leukemia on an annual basis for first ade of life (ICRP 1962; Sowby 1965; UNSCEAR 1962, 1964). As with chemical carcinogenesis risk assessment, therefore, the foundations of the LNT modeling for ing radiation-increased cancer risks are directly traced back to Lea, Timofeeff-Ressovsky et al. and ultimately to ler's proportionality rule. Chemical carcinogens Five years after the publication of the BEAR I report, Mantel and Bryan ( 196!) published their influential paper entitled "Safety' Testing of Carcinogenic Agents" based on the probit dose-response model in order to estimate tumor incidence for carcinogens. Biostatistical estimates of cer risks were first provided by Bryan and Shimkin (1943) when they applied the probit model to estimate the cancer risk of three carcinogenic hydrocarbons (i.e.,

cholanthrene; I ,2,5,6-dibenzanthracene; 3,4-benzpyrene) in strain C 3 H male mice. The motivation for Mantel and Bryan to develop the biostatistical model for predicting carcinogen risk was due to the fact that Mantel, a biostatistician at the US National Cancer Institute (NCI), was asked by the Director of the NCI to develop guidelines for the number of laboratory animals that would be needed to establish the safety of a test agent within the context of a hazard assessment.

This response followed a request, after the Thanksgiving berry scare of I959, by the Secretary of the Department of Health, Education and Welfare (HEW) to the NCI. The cranberry scare was a public relations nightmare in which trace residues of a cancer-causing herbicide

[i.e., amitrole (3-amino-1

,2,4-triazole)]

were detected in some sources of cranberries just before the holiday. The secretary of HEW recommended against buying cranberries that year, ing to a consumer panic that threatened the industry.

In order to avoid such situations in the future, the secretary of HEW requested the NCI to provide guidance on which cancer-causing substances were "safe" and at what dosage levels. Mantel and Bryan ( 1961) noted the generality of their modeling approach and proposed the concept of a virtually safe dose with an estimated risk of 1/100 million. Some 12 years later, the FDA would propose the use of the tel-Bryan

(!961) model and recommend the 1/100 million safety guide in their July I9, I973 risk assessment proposal in the Federal Register.

When the rule was finalized in 1977, the Mantel-Bryan probit model was retained but with several modifications and with the acceptable (de minimus) risk being reduced to 1/million.

This value was considered as the level below which no additional regulatory action would be taken within the context of the safety of animal carcinogen residues.

The finalized Mantel-Bryan model of the FDA was the first quantitative risk assessment model approved by a regulatory agency. Two years later, the FDA ( 1979) significantly revised the cancer risk assessment policy, replacing the modified Mantel-Bryan model with a linear dose-response model based on multiple factors, including its more conservative risk estimation and ease of calculations (Anonymous, ! 979). In the low-dose zone, the one-hit model discussed above is closely approximated by a simple linear model. The US EPA strategy for assessment and regulation of carcinogens displayed a profound evolution during the 1970s. Based on expert testimony during pesticide ings, EPA attorneys developed a legal brief that embodied "cancer principles" (NAS 1983). These "principles" gested that carcinogen exposures should be prevented.

As the concept of "banning" carcinogenic agents was soon seen as unrealistic, EPA quickly adopted non-regulatory guidelines for a general risk assessment process (EPA 1976). This process advocated the use of quantitative risk assessment as a means to differentiate risks among cals and engineering processes.

The guidance was very general, being limited to less than a page within the eral Register.

These guidelines were followed by a paper from the EPA Carcinogen Assessment Group (CAG) (Albert et a!. !977), which provided a strong endorsement of the LNT concept, arguing that linearity was supported by human epidemiological studies (e.g., ionizing radiation and cigarette smokiPg related lung cancer) and ity studies that were also claimed to follow a linear dose response and believed to be the underlying mechanisms of carcinogenesis.

In a March 15, 1979, Federal Register, the EPA Administrator Douglas Castle stated that "Risk ment from animal data is performed using the 'one-hit' model" based on the 1976 Interim Guidelines (EPA 1976). He went on to state that "the one-hit model was endorsed by the four agencies in the Interagency Regulatory son Group" based on its highly conservative nature and the uncertainties in extrapolating from animal data to human responses and the possibility that humans may be more ceptible than the animal model, because of broad human interindividual variability in exposures and "other unknown factors".

The strongly clarifying and underlying statement of the administrator was due in part to the fact that EPA had used other cancer risk assessment models under other latory acts and by other US federal agencies.

According to Albert ( 1994), Chair of the EPA cer Assessment Group (CAG) during the 1970s, the EPA Arch Toxicol adopted the linear no threshold model (LNT) of the Atomic Energy Commission (AEC) that had been applied to mating risks from fallout from atomic weapon tests. The LNT model was attractive to EPA since it was very simple to apply; all that was needed in a toxicological sense was to identify the lowest dose of agent that induced a statistically significant response and draw a straight line to the origin of the graph for the dose versus cancer incidence.

Its cal plausibility was based on the linearity of mutation dose response within the framework of target theory. He noted that "any difference between chemical carcinogens and ionizing radiation could be waived aside as they both cause genetic damage ... " Statisticians would argue that the straight line lation to zero from the lowest statistically significant response ignored data at the high doses. Thus, during a meeting of leading statisticians called by the CAG, a sion was made to change from the single-hit model to the multi-stage model since it used all the data, while ing linearity at low dose and being compatible with the concept of cancer being a multi-stage process. Consistent with this assessment, the NAS Safe Drinking Water mittee ( 1977) recommended the adoption of LNT modeling for risk assessment using a multi-stage model. However, in 1982, the Safe Drinking Water Committee (SDWC) was skeptical about LNT modeling for chemicals and rescinded its endorsement of the LNT model noting " ... more dence could be placed in mathematical models for olation if they incorporated biological characteristics of the animal studies ... since the users of this volume will be likely to favor different varieties of the conventional extrapolation models or will have access to some of the newer developmental methodologies, it is premature at this stage to recommend any single approach by selecting it for calculations

... " (p 8). However, since LNT modeling was already in use by EPA, in 1983, the SDWC again endorsed the LNT model and its subsequent use became the default methodology for chemical cancer risk assessment. ing to Albert ( 1994 ), none of the possible models (single hit, multi-hit, logit, probit, multi-stage, others) were ically credible.

The agency simply needed one that would be acceptable.

The agency applied LNT risk assessment methods using the multi-stage model for the regulation of trihalomethanes in drinking water in a November 29, 1979, notice in the Federal Register (EPA Environmental tion Agency (US EPA) 1979a, b), a process that would be followed in subsequent EPA cancer risk assessments.

The parallel, yet converging linear dose-response gies of the EPA and FDA represent the regulatory origin of current cancer risk assessment practices throughout the world. They are directly traced back to the efforts of Lea (I 946) and Timofeeff-Ressovsky et a!. ( 19:15), all of which stemmed from the "Proportionality Rule" of Muller (1930).

Arch Toxicol Eugenics While the LNT concept for mutation was born within the intellectual and scientific framework of the physics-based radiation target theory, its applications also found portive resonance within the philosophical, ideological and political frameworks of eugenics.

German eugenicists expressed considerable concern that ionizing radiation may hurt the German germ plasm (Proctor !999; Martius 1931). Educational programs based on these concerns cautioned against exposures to ionizing radiation that might adversely affect future generations of Germans. Recommendations as early as 1927 by the Bavarian Society for Pediatrics and Gynecology stated that women receiving excess X-rays during pregnancy should abort their fetuses. Pushing this concept even further, in 1930, Eugene Fisher, director of the Kaiser Wilhelm Institute for Anthropology, argued that women exposed to X-rays should be permanently vented from having children (Proctor 1999). Muller's own history is replete with his highly visible association with national and international activities advancing eugenics philosophy and agenda. Even as late as 1955, Muller gave a strong eugenics advocacy presentation in Germany, ing such ideas with a large audience of Nobel Prize winners (The Lindau Mediatheque

!955). The biophysical concept of the gene had important eugenics implications.

Since mutations could be induced by ionizing radiation in a linear at low-dose manner, this concept provided the principal foundation that all ing radiation-whether via medical diagnosis/treatment or industrially-was a concern for "genetic health". The genetic toxicology studies of Timofeeff-Ressovsky et al. (! 935) transformed these above-cited radiation health cerns, providing biophysical models and the LNT-single-hit model risk assessment paradigm.

Such actions provided a key vehicle by which eugenics would focus on tion protection for preventing the occurrence of genetic defects. In fact, the development and activities of the ics department of the Kaiser Willheim Institute under the direction ofTimofeeff-Ressovsky was affected by such spectives (Gausemeier 20 I 0). The concept of LNT for ionizing radiation-induced mutation was, therefore, built upon a scientific/cultural framework and applied to a range of health-related cies, especially those of eugenics during the early ades after the discovery of X-ray-induced mutations.

In fact, the eugenics area would serve as an intellectual training ground for how ideas such as LNT could be "softened", humanized and successfully integrated within a post-World War II society. Some aspects of eugenics advocacy and the LNT concept would morph into ern regulatory policy for carcinogen regulation, evolving from that of preserving the gene pool of certain racial subgroups or other targeted populations to a humanistic framework that would reduce mutational risks to entire populations.

Evolution and endogenous mutations The LNT had its start in an attempt to explain evolution, finding other outlets in the world of eugenics and later public health regulatory policies.

While Muller was a leader in these activities, he did not abandon his quest to determine those underlying factors that served to provide the novel mutations for natural selection.

In fact, prior to his discovery of X-ray-induced mutations in 1927, Muller reported that temperature increases enhanced the mutation rate by about two-fold (Muller !921-\c).

However, the temperature hypothesis was placed on the research back burner when high doses of X-rays were found to markedly enhance mutation frequency.

Muller would return to the temperature-evolution hypothesis some three decades later, completing an intellectual and professional circle, reflected in the comments of Plough and Ives ( !934), his former colleagues at Amherst lege (1940-1945) who noted that "since Muller and Mott-Smith conclude that natural radiation is inadequate to account for mutations in nature, it seems possible to suggest that ubiquitous temperature variations may play that role". If Muller had lived into the decades of the 1980s (he died in the 1967), he would have begun to appreciate the so-called other conditions suggested by Oliver ( !93 l) as the cause of the overwhelming portion of spontaneously occurring mutations is now believed to be derived from endogenous metabolism, for which complex and integrative DNA repair processes have been selected for via natural selection (De Bont and van Larebeke 200-l; Lindahl 1996). Summary The LNT concept was initially proposed to account for evolutionary change and then later applied for the ment of risks for some genetic diseases and cancer dence (Table 2). The initial data upon which the LNT concept was based were limited to a few studies of an acute nature and at very high doses. Within a decade, the LNT dose-response model was provided with a nistic foundation via the integration of the single-hit concept within target theory. The LNT-single-hit model was then used by radiation geneticists to frame the lectual debate on low-dose ionizing radiation risk to the human genome. It provided the basis for the dations of the US NAS BEAR I Committee in 1956 for Arch Toxicol Table 2 LNT history: the temporal sequence leading to the LNT dose-response model for cancer risk assessment References Muller ( 1927) Olson and Lewis ( 1928) Muller ( 1930) Timofeeff-Ressovsky et al. (I BEAR I 1956 (Biological Effects of Atomic Radiation Committee, Genetics Panel) Mantel and Bryan ( l% I) FDA (!973) EPA ( 197(i) (see Albert et al. ( 1977), Anonymous ( 1979) FDA (1977) U.S. NAS Safe Drinking Water Committee ( 1977) FDA (1979) EPA (l979a, b) Specific temporal events Mutation findings-X-rays induce mutations in fruit flies .JJ. LNT model proposed to account for evolutionary changes following Muller's discovery that X-rays can induce mutations in fruit fly germ cells .JJ. Develops proportionality rule (i.e., linear dose response) for ionizing radiation-induced genicity .JJ. Application of radiation target theory for mutagens.

Used target theory to propose a hit theory for ionizing radiation-induced mutation.

The hit mechanism was used to explain the LNT dose response .JJ. Proposes the use of the linear dose-response model for germ cell mutation, using the "doubling rule" .JJ. Develops carcinogen risk assessment model based on the probit model. This activity was undertaken to advise US governmental agencies on chemical risk assessment .JJ. Proposes a probit-based quantitative risk assessment method for cancer risk based on the tel and Bryan 1961 paper. The proposal stated that an acceptable risk was 1/100 million .JJ. Proposed guidelines for carcinogen risk assessment based on quantitative risk assessment.

Recommended a linear dose-response model .JJ. FDA rule finalized, retaining the Mantel-Bryan model with some modifications.

The acceptable risk value was changed to Ill million (10-6) .JJ. Recommended that EPA adopt LNT for carcinogen risk assessment.

This recommendation was profoundly significant given the widespread multimedia regulatory functions of EPA. Within 2 years of the recommendation, EPA applied the LNT to the regulations of trihalomethanes (e.g., chloroform) in drinking water .JJ. Replaced the modified Mantel-Bryan model with the LNT model for carcinogen risk ment, based on the following reasons: I. Linear procedure is least likely to underestimate risk. 2. Linear extrapolation does not require complicated mathematical procedures.

3. No arbitrary slope is needed to carry out linear extrapolation.

4. Several significant limitations were found with the application of the Mantel-Bryan model (Anonymous 1 979) .JJ. EPA established a national drinking water standard for trihalomethanes (including chloroform) based on an LNT methodology as recommended by the US NAS Safe Drinking Water mittee ( l 977) the switch from a threshold to a linear dose-response model for estimating ionizing radiation-induced germ cell mutation using the doubling dose concept. The single-hit model was soon generalized to the process of cancer risk assessment and adopted by national and national committees concerned with ionizing radiation by the late 1950s and early 1960s. Five years later, Mantel and Bryan ( 1961 ), researchers at the US National cer Institute, proposed a probit model-based cancer risk assessment method. It was the Mantel and Bryan ( 196 t) model that was proposed by the FDA in 1973 for cer risk assessment procedures, being replaced with a LNT model by the FDA in 1979, the same year that EPA applied the LNT for the regulation of carcinogens (i.e., trihalomethanes) in drinking water. The LNT model and its single-hit explanation/mechanism theory, therefore, can be traced back to the concept of radiation-induced mutation target theory as proposed by vsky et a!. ( 1935), which was founded on the ality rule of Muller ( 19 JO) which itself had its origins in the 1928 paper of Olson and Gilbert that created the LNT concept following the seminal findings of Muller ( 1927) that ionizing radiation could induce mutation in the germ cells of fruit flies.

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new findings challenge 4 historical foundations of the linear dose response 5 Edward J, Calabrese 6 Received:

24 April2013 I Accepted:

II July 2013 7 ©Springer-Verlag Berlin Heidelberg 2013 a Abstract This paper extends several recent publi-9 cations indicating that Hermann J. Muller: (1) Made 10 deceptive statements during his Noble Prize Lecture 11 on December 12, 1946, that were intended to promote 12 the acceptance of the linear dose-response model for 13 risk assessment for ionizing radiation and (2) that such 14 actions of Muller were masked by a series of decisions 15 by Muller's long-time colleague and esteemed radia-16 tion geneticist Curt Stern, affecting key publications in 17 the mutation literature.

Such actions further enhanced 18 acceptance of the linearity dose-response model while 19 preventing Muller's deceptions from being discovered.

20 This paper provides documentation that Muller rein-21 forced such practices within the scientific literature in 22 the early 1950s, by supporting scientifically question-23 able actions of Stern. Detailed documentation is provided 24 that demonstrates how these actions affected national and 25 international risk assessment policy for ionizing radiation 26 and chemical carcinogens via the recommendations of 27 the National Academy of Sciences Biological Effects of 28 Atomic Radiation committee in 1956, to adopt the linear 29 dose-response model. 30 Keywords Mutation

• Linearity

• Dose response

• Risk 31 assessment

• History of science

• Muller A 1 E. J. Calabrese (121) A2 Department of Public Health, Environmental Health Sciences, A3 University of Massachusetts, Morrill I, N344, Amherst, MA A4 01003, USA A5 e-mail: edwardc@schoolph.umass.edu J oumal : Large 204 Article No : 1105 MS Code : 1105 Introduction It was recently discovered that the 1946 Nobel Prize ture for Biology and Medicine by Laureate Hermann J. Muller misled the audience on the nature of the dose response in the low-dose zone concerning the effects of ionizing radiation on germ-cell mutagenicity to advance an ideologically motivated risk assessment policy brese 20 l I a, b, 20! 2). Evidence to support this sion is found in Muller's own words from letters he sent to Professor Curt Stern of the University of Rochester, an expert in radiation genetics.

Stern sent Muller a script by Ernst Caspari and himself on November 6, 1946, for review as Muller was a paid consultant to the project (Calabrese 20 I I c). This manuscript demonstrated port for a threshold dose response, while challenging the linear dose-response single-hit mutagenicity mechanism model, based on an extensive study of ionizing radiation on mutation in the germ cells of male fruit flies. On ber 12, 1946, Muller acknowledged receipt, noting that the findings strongly challenged the linearity dose-response concept and, given their importance, needed to be licated as soon as possible (Calabrese 20 II c). This term study used the lowest ionizing radiation dose rate yet reported.

Despite this new information, Muller would go on to deliver his Nobel Prize Lecture some 5 weeks later (December 12, 1946), proclaiming that one could no longer consider the possibility of a threshold dose response for germ-cell mutagenicity.

The only option, he argued, was to switch to a linearity dose-response model for risk ment (Muller l946a). Muller, of course, made these public claims while ing that the most extensive and relevant testing supported a threshold interpretation.

A letter from Muller to Stern 5 weeks after the Nobel Prize Lecture (January 14, 1947) Dispatch : 1-8-2013 0 LE li1l CP Pages : 19 0 TYPESET li1l DISK 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 66 confirmed his support for study replication, that he had no 67 technical criticisms of the Caspari study, and supported 68 publication especially in view of the caveats worked into 69 the discussion, hopefully preventing acceptance of a thresh-70 old interpretation (Calabrese 20 12; Lilly Library 1947 a, 71 January 14 Jetter). In effect, Muller told the Nobel Prize 72 Lecture audience one story while in private correspond-73 ence he revealed a profoundly different view. According to 74 his former student, friend, and colleague, Crow (1995), it 75 was well known that Muller would try to win arguments by 76 exaggeration and overstatement.

Crow found this behavior 77 exasperating as Muller would often end up hurting his case 78 by unnecessarily misrepresenting facts and circumstances, 79 incorrectly thinking it would help him win his argument.

80 This same behavioral trait was evident at the Nobel Prize 81 Lecture. 82 Before his Nobel Prize Lecture, Muller sought to 83 raise concern over the public health implications of ion-84 izing radiation and to change the risk assessment process 85 for ionizing radiation from the use of a threshold dose-86 response model to the far more conservative linear dose 87 response.

This goal was essentially shared by the entire 88 radiation geneticist community.

Following his Lecture, 89 Muller would now have two goals: Protecting his reputa-90 tion by ensuring that his misleading comments would not 91 be discovered while still aggressively pushing acceptance 92 of the linearity agenda. Both goals were entangled; being 93 such an important scientist and leader any fall in Muller's 94 status would have a devastating impact on the acceptance 95 of the linearity dose response, especially if it involved an 96 ideological misrepresentation about the linearity concept. 97 Muller achieved both goals due to decisions of Stern that 98 discredited the findings of his colleague and co-author 99 Ernst Caspari, thus saving Muller from criticisms about 100 his Nobel Prize Lecture while supporting the question-101 able findings of Delta Uphoff, another co-author.

Mul-102 ler's misleading comments and the Stem's apparent data 103 obfuscations would not be revealed for more than 60 years 104 while the linearity acceptance goal by regulatory agencies 105 worldwide was attained.

The present paper extends the 106 recent reports of Calabrese (20! l a, b, 2012) with newly 107 discovered findings that demonstrate a carefully focused 108 and timed set of inexplicable scientific judgments by Mul-109 ler concerning the nature of the dose response.

These 110 actions reinforced his Nobel Prize Lecture and 111 the actions of Stern that enhanced the goal of achieving 112 a switch from threshold to linearity.

This paper also dem-113 onstrates the profound impact of the Stem/Muller actions 114 on the radiation genetics community based on the scien-115 tific publication record and dose-response recommenda-116 tions/conclusions supporting a linearity dose-response risk 117 assessment model by the highly influential NAS BEAR I 118 Committee, Genetics Panel.

Journal : Large 204 Article No : 1105 MS Code : 1105 Arch Texico! Part 1-Stern's plan to promote linearity 119 Curt Stem was a long-time supporter of the idea that ion-120 izing radiation affected germ-cell mutation in a linear dose-121 response manner. He expected that this would be observed 122 in studies he was directing under the aegis of the Manhat-123 tan Project using fruit flies. While a linearity dose-response 124 was reported in acute studies with X-rays (Spencer and 125 Stem 1948), the most significant test would take place 126 with the research of Ernst Caspari when gamma radiation 127 would be administered up to a 13,200-fold lower rate than 128 in the Spencer research.

In a troubling development, Cas-129 pari reported to Stem that his findings did not support a 130 linear interpretation but rather a threshold dose response.

131 Based on Jetter correspondence between Stem and Caspari, 132 Stem initially refused to accept this interpretation, arguing 133 that the mutation threshold response was most likely due 134 to unusually high control group values (i.e., spontaneous 135 mutations in sperm stored in the spermatheca of the female 136 for 3 weeks) which masked a radiation-induced treatment 137 effect (Calabrese 20 l I b). Cas pari then researched this 138 issue by exploring the literature and obtaining substantial 139 unpublished data on this specific issue from Muller based 140 on research during his appointment at Amherst College 141 (1940-1945).

Caspari argued that his control group muta-142 tion data were not aberrant but consistent with the litera-143 ture and Muller's data for aged sperm whether stored in 144 the spermatheca of the female or in the male. As a result 145 of the Caspari analysis, Stern withdrew his objection and 146 accepted the conclusion that the control group spontaneous 147 mutation values were within the normal range. Since Stem 148 could not dismiss the findings of Caspari due to the con-149 trois, he then opted for an alternative but bizarre strategy to 150 marginalize the threshold dose-response conclusion.

Stem 151 directed the manuscript discussion to explain why these 152 data should not be accepted and utilized until it was deter-153 mined why Caspari's findings differed from those of Spen-154 cer and Stem's acute study which they claimed supported 155 linearity.

It was this manuscript of Caspari that was sent to 156 Muller for review just prior to his Noble Prize Lecture. 157 It is odd that investigators reporting on striking new 158 findings, using the most advanced methods and the low-159 est dose rate yet studied, would demand the reader not take 160 the data seriously.

Stem placed no such restriction upon the 161 Spencer paper, a study with considerable methodological 162 limitations

[e.g., inadequate control groups, inappropriate 163 data combining for statistical analysis, lack of adequate 164 X-ray instrumentation calibration, poor temperature con-165 trol, and dose rates differing by as much as 10-fold (10 166 and 100 r/min) between treatments, thereby creating two 167 experimental variables within one experiment] (Calabrese 168 201! b). Furthermore, there were at least two dozen signifi-169 cant methodological differences between the two studies 170 Dispatch : 1-8-2013 0 LE li:l CP Pages : 19 0 TYPESET li:l DISK Arch Toxicol 111 making them not directly comparable.

Stern published the 112 manuscript (Caspari and Stern 1948) with its misdirected 173 discussion, without apparent independent, peer review in 174 the journal for which he was the editor, that is, Genetics.

175 Comment 176 Based on this temporal sequence, it would appear that the 177 principal driving force to challenge the Caspari findings 178 that supported a threshold interpretation was his advisor 179 and co-author, Curt Stern. It was Muller who indicated 180 that the findings of Caspari needed to be replicated since 181 they were contrary to a linear single-hit dose-response 182 interpretation.

Of particular note, however, was that the 183 only changes made to the Caspari manuscript following 184 the review of Muller was to add the name of Muller to the 185 acknowledgments section and to remove the statement 186 from the conclusion that the findings supported a tolerance 187 or threshold interpretation (Calabrese 20 l I b). 188 Part 2-the replication studies 189 Since Ernst Caspari and Warren Spencer were no longer 190 available to continue experimentation, Stern engaged the 191 services of a Master's student, Delta Uphoff, to assess why 192 the Caspari study did not support a linear interpretation.

193 The results of the initial experiment were deemed by Stern 194 as not usable as her control group spontaneous mutation 195 rate was strikingly low, being outside the expected range 196 for aged sperm ( -40 % lower than expected);

no conclu-197 sions could be drawn from the study (Uphoff and Stern 198 1947). A similar very low control group spontaneous muta-199 tion rate response for aged sperm in her second experi-200 ment would also make such data uninterruptable.

In her 201 third and final experiment, Uphoff reported control values 202 in the normal range for aged sperm but the radiation treat-203 ment response was itself aberrant, far exceeding predicted 204 responses assuming low-dose linearity (Calabrese 20 l ! b). 205 Stern: What to do next 206 Finding a way to support linearity was the prevailing 207 theme. For example, when Caspari had shared his data with 208 the Head of Genetics at the Brookhaven National Labora-209 tory and future member of the BEAR I Committee/Genet-210 ics Panel, Milislav Demerec, he wrote to Caspari asking 211 what can be done to save the single "hit" linearity dose-212 response paradigm (Calabrese 20 I I b; American Philosoph-213 ical Society 19471', September 25). The "hit theory" for 214 ionizing radiation-induced mutation was first postulated by 215 Timofeeff-Ressovsky et al. (! 935), providing a theoretical Journal : Large 204 Article No : 1105 MS Code : 1105 mechanistic foundation for the LNT dose-response model. 216 Given his goals and ideology, Stern had little choice. 217 Another experiment was not going to be practical as Uphoff 218 would leave for a position with the NIH. In the absence of 219 new data, Stern decided upon a new strategy to "save" the 220 single-hit linearity dose response.

In order to achieve this 221 goal, he would have to do two things: (1) Reverse his posi-222 tion on the Uphoff control group data, declare that they 223 are normal, not aberrant, making the Uphoff experiments 224 now interpretable and (2) challenge further the credibil-225 ity and acceptance of the Caspari study (i.e., beyond the 226 misdirected discussion of the Caspari!Stern paper). Stern 227 took the bold action of asserting that the Uphoff control 228 group data were part of the normal distribution.

He offered 229 no explanation or assessment of the literature to justify 230 this conclusion.

This would not be difficult as only very 231 few people would have known about his earlier concerns 232 with the Uphoff control group data, since the manuscript 233 (Uphoff and Stern 1947) detailing such concerns was never 234 submitted for publication but was placed in the Atomic 235 Energy Commission (AEC) archives, initially as a classi-236 fied manuscript.

Thus, the written critique of the Uphoff 237 control group data and letter communications on this topic 238 were generally not known or available.

239 The Uphoff and Stern (I 949) paper also raised anum-240 ber of doubts about the Caspari paper such as whether its 241 non-treatment effect/threshold finding was the result of 242 "errors in sampling." Given standard professional proto-243 col, the "errors in sampling" hypothesis was a surprising 244 and unexpectedly harsh challenge to the work of Caspari, 245 a University of Rochester team member, especially since 246 this criticism had never been raised previously by Stern, 247 Muller, or others in previous detailed evaluations.

In fact, 248 there was never any documentation to support this possi-249 bility. Further, Stern also raised the specter of the Caspari 250 control being elevated by unnecessarily stating that his 251 control group was higher than each of the controls of the 252 three Uphoff experiments.

Stern neglected to state that two 253 of the Uphoff studies had aberrantly low control group val-254 ues based on the published literature and Muller's data. 255 This decision by Stern would now make the Uphoff experi-256 mental data "interpretable," whereas several months before 257 he judged it as "uninterpretable." Also, the third Uphoff 258 experimental control data were indistinguishable statisti-259 cally from the Caspari control (0.2489 vs. 0.2352 %). Such 260 actions helped to achieve the above-stated goals of enhanc-261 ing the credibility of the Uphoff data while marginalizing 262 the Caspari findings.

263 The Uphoff and Stern ( 1 949) paper changed the way the 264 Caspari data (Caspari and Stern l94R) were perceived and 265 accepted by members of the scientific community.

Below 266 are quotes from several papers (Higgins 195!; Singleton 267 l954a, b) and a dissertation (Jolly 2004) that address very 268 Dispatch : 1*8-2013 0 LE li1l CP Pages : 19 0 TYPESET li1l DISK 269 clearly how the Uphoff and Stern ( 1949) paper marginal-270 ized the research of Caspari. Of particular significance is 271 that the judgments drawn by each of these papers were fac-272 tually and interpretationally incorrect.

273 Higgins ( 195!) stated that "Uphoff and Stern 274 (! 949) ... concluded that low-level radiation does produce 275 mutations in fruit-fly sperm and that the apparent inconsist-276 encies of previous results were due to different experimen-277 tal techniques and errors in sampling" (page 10, column 1). 278 Singleton ( 1954a) stated that "Cas pari and Stern ( !948) 279 studying chronic gamma radiation found no increase over 280 controls for doses of 2.5 r/day for 21 days. However, it was 281 later documented by Uphoff and Stern (

that the con-282 trois used by Caspari and Stern had an abnormally high sex 283 linked lethal frequency and that actually there was an effect 284 of the chronic gamma radiation of 2.5 r/day." (page 599) 285 Jolly (2004) stated (1) that "Stern and Cas pari initially 286 detected no significant difference in the mutation rates on 287 the controls and the irradiated flies, though later they cor-288 rected for experimental errors and got a statistically signifi-289 cant difference." (pages 78-79) (2) "The results of Stern's 290 initial experiment failed to support the linear hypothesis for 291 genetic injury. Assuming that something must have been 292 wrong with the experiment, he eventually identified experi-293 mental errors, which, when corrected for, supported linear-294 ity." (pages 80-81). 295 Caspari's control group data were therefore once again 296 challenged by Stern; the once aberrantly low controls 297 of Uphoff were now seen as being in the normal range. 298 With these changes, the dose response of the collective 299 grouping of the Stern Drosophila experiments would 300 appear linear. This is the conclusion of what Uphoff 301 and Stern published in their one-page technical note in 302 the 1949 Science article summarizing the Spencer and 303 Stern (

and Cas pari and Stern ( 1941\) papers and 304 the three Uphoff experiments.

This 1949 paper, as noted 305 above, did not include mention that the previous conclu-306 sions (Uphoff and Stern 1947) about the Caspari and the 307 Uphoff control groups that had been reversed by Stern 308 and the role of the Muller data assessment in the deci-309 sion-making process. Since the Uphoff and Stern (! 949) 310 brief technical paper lacked any information on research 311 methods and other relevant data, the authors promised 312 a detailed follow-up publication to correct this critical 313 limitation, a promise never fulfilled.

Given the lack of 314 information provided in the Science paper and the pres-315 tige of this journal, it raises a question about the circum-316 stances surrounding its publication within this context. 317 It should be noted that Hermann J. Muller's first gradu-318 ate student (i.e .. H. Bentley Glass) became an editor at 319 Science in 1948, only months prior to the submission 320 of the Uphoff and Stern manuscript.

Glass also had a Journal : Large 204 Article No : 1105 MS Code : 11 OS Arch Toxicol relationship with Stern with whom he had been awarded 321 a National Research Council post-doctoral fellowship at 322 the Kaiser Wilhelm Institute in Berlin (Erk 2009). Since 323 Glass was an expert on Drosophila radiation genetics, it 324 is likely that he oversaw the evaluation of the manuscript.

325 One must also question to what extent Muller/Stern may 326 have exploited their relationship with Glass to facilitate 327 the publication of such a limited paper and used the jour-328 nal to advance an ideological perspective.

329 Muller's post Nobel Prize dose-response comments 330 about the Caspari and Stern (1948) study 331 Muller's statement 332 In his 1950 article entitled "Some present problems in 333 the genetic effects of radiation" in the Journal of Cellular 334 and Comparative Physiology Muller ( l950a) provided an 335 explicit characterization of the Cas pari and Stern ( 1948) 336 findings.

Muller stated on page I 0 "A recent paper by 337 Spencer and Stern ....... extends the principle (i.e., one-338 hit principle) down to total doses of 50 r and 25 r." In the 339 next paragraph, he stated: "It is true, in a parallel paper... 340 .Caspari and Stern have reported results somewhat deviat-341 ing from the above." 342 Comment 343 Muller trivialized the significant challenge of the Caspari 344 study to the linearity dose-response paradigm.

The key 345 Muller phase concerning the Caspari data is "somewhat 346 deviating".

The Spencer and Stern ( 19-1-8) study involved an 347 acute exposure, that is, all doses of radiation were admin-348 istered within a few minutes to a few hours. In contrast, 349 the .Caspari and Stern (1948) study provided the same 350 total dose as in the Spencer and Stern study but spread 351 over 21 days, at a dose rate up to 13,200-fold lower. The 352 "somewhat deviating" results were such that at the lower 353 dose rate of the Caspari and Stern study, the data supported 354 a threshold interpretation, not the expected linear propor-355 tionality response.

Muller was quite concerned with the 356 Caspari study as it represented a potentially significant 357 challenge to linearity, repeating this perspective in letters 358 (Lilly Library 1947a, January 14; American Philosophi-359 cal Society 1946, November 12) to Stern and emphasizing 360 the need to replicate this study, despite the requirement for 361 additional funding and the efforts of multiple scientists and 362 staff for about 1 year. It is also important to note that Mul-363 ler never mentioned any of the numerous methodological/

364 analysis limitations/flaws of the Spencer and Stern ( 1948) 365 in any of his publications.

366 Dispatch : 1-8-2013 0 LE 0 CP Pages : 19 0 TYPESET 0 DISK 367 368 369 370 371 372 373 374 375 376 377 378 379 380 381 382 383 384 385 386 387 388 389 390 391 392 393 394 395 396 397 398 399 400 401 402 403 404 405 406 407 408 409 410 411 412 413 414 415 416 Arch Toxicol Muller's statement In footnote I on page 10 of the above-cited article, Mul-ler ( 1950a) stated that "Uphoff and Stem have published a report of further work, with doses as low as 50 r, given an intensity as low as 0.0165 r per minute. The results obtained are entirely in conformity with the one-hit prin-ciple. A consideration of these results, together with the early work, leads to the conclusion that the deviation first referred to (the Caspari and Stem l 94R findings) was caused by a value for spontaneous mutation rate that hap-pened to be unusually high." Comments Muller claims that the research of Delta Uphoff and Curt Stem is "entirely in conformity with the one-hit principle" (Timofeeff-Ressovsky et a!. l9:i5). What Muller neglected to state was: (1) Uphoff's first experiment displayed an aberrantly low control group response based on Muller's own extensive data involving some 200,000 fruit flies (Muller l946b ). A letter from Curt Stem to Ernst Cas pari (undated) (American Philosophical Society Undated, circa July-Aug 1947) addressed the control group issue. It states: "The radiation data continues to be puzzling.

Delta's dif-ference between control and exper[imental group] appears to be due mainly to a much lower control group value than yours. However, Muller informs me that his data give an aged control value close to yours. Thus, my first idea that your results could be "explained away" by assuming that your control value happened to be unusually high, seems unlikely.

Rather does Delta's control appear too low. Well, we'll have to meet." Muller provided this information to Stem twice in letters dated February 3, 1947, and August 4, 1947 (Lilly Library l947b, c). It should be noted that the occurrence of increased mutations in aged sperm in the control group as reported by Caspari was not a new concept to Stem. In fact, when Timofeeff-Ressovsky first presented such data in the late 1930s, Stem corresponded with Demerec specifically addressing these findings.

These letter exchanges reveal not only Stem's knowledge of the findings, but also of his knowledge that the findings had been subsequently replicated (Lilly Library 1938a, b, c). The report of Rajewski and Timofeeff-Ressovsky (I 939) on this topic would most likely have considerable scientific weight as Timofeeff-Ressovsky was on par with Muller for scientific reputation in the area of radiation genetics.

In the Atomic Energy Commission (AEC) manuscript by Uphoff and Stem ( 1947) concerning her replication of the Caspari study, the low response control group issue was explicitly addressed as follows in their "Discussion" section. "In his extensive studies on the effect of aging on the mutation rate in sperm, H.J. Muller (unpublished) has Journal : Large 204 Article No : 1105 MS Code : 11 OS found a weekly increase of about 0.07 % for sex-linked lethals in various stocks kept at 25 °C. At 18 °C, the temper-ature used for aging in the laboratory, the weekly increases may be assumed to be slightly less, perhaps 0.05 %. Taking a value of 0.10 %, similar to that of Spencer and Stem's control rate, for sperm before aging, the expected control rate after aging should be approximately 0.25 %. This fig-ure is much closer to the control rate observed by Caspari and Stem than to that found in the present work." In their acknowledgments of this manuscript, Uphoff and Stem stated that "we are very grateful to Dr. H. J. Muller for his permission to quote from his unpublished data." Thus, Mul-ler would have known that his research was used to evalu-ate the reliability of the Caspari and Uphoff control groups. The control group response of Uphoff and Stem ( 194 7) was sufficiently low such that they stated that the data were uninterpretable (i.e., "a final interpretation of these results cannot be offered.").

Uphoff and Stem ( !94 7) explicitly raised the possibility that the low control group values "may reflect a personal bias of the experimenter." The man-uscript did not identify whether the bias concern statement was directed to Stem, Uphoff or both, or the type of bias. (2) Uphoff's second experiment also displayed a similarly aberrant low control group response, likewise affecting the possible utility of the data. (3) The third (and final) Uphoff experiment obtained control values in the normal range but an aberrantly high treatment response, even assuming a linearity dose response (see Calabrese 20! ! a for a detailed evaluation). "Appendix" section provides the temporal let-ter exchange between Stem and Muller on the key ques-tion of control group mutation frequency upon which the acceptance of the Cas pari and Uphoff studies are based. Muller ( l950b) discredits the conclusion of Cas pari and Stem ( 1948) by asserting that the control group val-ues were unusually high. (1) Muller failed to state that the "high" control value of Caspari and Stem ( 1948) was first put forward as a criticism by Stem in the fall of 1946, when Caspari informed Stem that his findings supported a threshold, rather than a linearity interpretation.

(2) He also did not report that Caspari successfully rebutted Stem by presenting data on control group responses from published studies in the literature and from unpublished data provided by Muller himself. Muller failed to state that he had pub-lished a summary of the *mutation rate of sperm stored in the spermatheca for several weeks (Muller 1945). This is the information that he sent to Stem that supported the reli-ability of the Caspari control group data and marginalized the Uphoff study control group (see "Appendix" section).

Later studies by Muller and his student Helen L. Byers at the University oflndiana also supported the Caspari control group mutation frequency (Byers 1954; Byers and Mul-ler l 952). Nonetheless, Muller ( l 954h) would inexplica-bly continue his criticism of the Caspari and Stem (194?\) Dispatch : 1-8-2013 0 LE lil CP Pages : 19 0 TYPESET lil DISK 417 418 419 420 421 422 423 424 425 426 427 428 429 430 431 432 433 434 435 436 437 438 439 440 441 442 443 444 445 446 447 448 449 450 451 452 453 454 455 456 457 458 459 460 461 462 463 464 465 466 467 468 469 470 study, repeating the "unusually high control frequency" 471 (page 476) conclusion as a basis to reject its challenge to 472 linearity.

The question may be raised as to why Muller 473 would directly contradict himself on such a serious mat-474 ter and never be exposed to criticism.

While any answers 475 to this question must be speculative, Sankaranarayanan 476 and Wassom (2008) unequivocally state that Muller was an 477 "unquestioned authority," suggesting that it would be quite 478 difficult to challenge him or even consider doing so. 479 It should be noted that in early 1949, Muller became 480 concerned that Robley Evans of MIT was publishing a 481 paper in the journal Science on the mutagenic effects of 482 ionizing radiation and the nature of the dose response in the 483 low-dose zone. Muller had reviewed the manuscript prior 484 to publication and was upset that Evans had given credibil-485 ity to the Caspari and Stem (

paper. Muller wrote to 486 Stem (Lilly Library I 949, February 5) requesting that Stern 487 contact Evans and try to convince Evans to withdraw his 488 support for the Cas pari and Stem (I 948) findings.

There 489 is no evidence that Stern did this based on correspondence 490 records. However, it is possible that the subsequent attack 491 of Muller (1950a, b) on the Caspari and Stem (1948) find-492 ings was stimulated by this Evans paper (! 949) which 493 would need to be "neutralized." 494 Muller (1 954b) also further criticized the Cas pari and 495 Stem ( 1948) paper in a vague manner as being "more 496 doubtful than the others on some other grounds" (page 497 476), which he never clarified.

Such criticism may have 498 referred to the fact that Uphoff and Stem ( 1947) introduced 499 a modified method of counting sex-linked recessive lethals, 500 one that was different than reported by Caspari and Stem 501 ( !948) and also different than Spencer and Stem ( 1911\). 502 Uphoff and Stem ( 1947) recounted (i.e., adjusted) the Cas-503 pari and Stern ( !948) data with the new counting method 504 in order for it to be as directly comparable to their study 505 as possible.

The results of those adjustments were deemed 506 by Uphoff and Stem to be insignificant in their 1947 paper, 507 resulting in control and treatment responses that were, in 508 fact, even more similar than before the adjustment (i.e., 509 without a treatment effect). The published paper of Caspari 510 and Stern ( l94S) did not incorporate this adjustment (per-511 haps resulting in the veiled criticism of Muller 195-ta, b), 512 whereas the Uphoff and Stern ( !94 7) manuscript presented 513 the original and adjusted data; only these adjusted data 514 were used for the Cas pari and Stern ( !94S) data as sum-515 marized in the 1949 paper in Science by Uphoff and Stem. 516 Regardless, the adjustment for differing lethality estimation 517 techniques did not affect the study interpretation.

In a letter 518 on February 9, 1949, to Caspari in anticipation of the Sci-519 ence publication, Stem (American Philosophical Society 520 1949, February 9) stated that "It will be shown below (the 521 Science manuscript) that the difference in defining a lethal 522 is of no significance in the evaluation of the results."

Journal : Large 204 Article No : 1105 MS Code : 1105 Arch Taxi col In his 1950 papers, Muller never addressed any of these 523 critical issues that might affect a decision on the nature of 524 the dose response (Muller l950a, b). He also failed to state 525 that the Uphoff and Stem (1949) paper was only a one-page 526 summary, has very low control group values, no presenta-527 tion of research methods and that Uphoff and Stem ( !949) 528 promised to publish a detailed paper with all the missing 529 methods and data but had not (and never did). By discredit-530 ing the Caspari and Stem ( 1948) paper and restoring the 531 Uphoff data, Muller was able to protect his scientific repu-532 tation, his ethical standing and to give strong support to the 533 linearity single-hit theory dose-response model. 534 In a second paper in 1950 entitled Radiation Damage 535 to the Genetic Material in the American Scientist, Mul-536 ler ( 1950b) used the findings of Stem and his colleagues 537 to extend "the principle of proportionality of mutation fre-538 quency to dose down to doses of 50 r and 25 r and of Jess 539 than 0.001 r per minute, with a time-intensity relation dif-540 fering by over 400,000 times from that of our high intensity 541 dose." 542 Comment 543 By using the now revitalized data of Uphoff, Muller made 544 the claim of linearity over a 400,000-fold dose range. This 545 was a major conclusion as it gave an assertion of linearity 546 at low dose by a Noble Prize winner who had great author-547 ity within the field. Furthermore, Stem ( 1960) continued to 548 affirm the findings of Uphoff and Stem ( 1949) in the sec-549 ond edition of his acclaimed genetics textbook, published 550 in English, German, Japanese, Polish, Russian, and Spanish 551 (American Philosophical Society 1973, November) (auto-552 biographical statement), by stating that the dose rate had no 553 impact on the mutation incidence in Drosophila, whether 554 administered acutely or given "slowly and continuously, that 555 is, 'chronically,'

given over a long period." In order for Stem 556 ( 1960) to have reached this conclusion, he had to diminish 557 the findings of Cas pari and Stem ( l94S) and accept those 558 of Uphoff and Stem ( 1949). A further note is that the Mul-559 ler ( l950b) paper contradicted his 1950a paper on the dose 560 rate: The two papers used a different lowest dose rate: 0.001 561 r/min (Muller l950b) versus 0.00165 r/min (50 r/30240 min 562 in 21 days) (Muller 1950a)-a 65-fold difference.

Muller 563 (1950b) rounded down the 0.00165 r/min rate to 0.001 r/ 564 min, increasing the extrapolation range from approximately 565 250,000-to 400,000-fold.

Why Muller rounded the num-566 bers down is not known, nor was it necessary.

Secondly, if 567 rounding was to occur it would normally have been rounded 568 up to 0.002 r/min. This action of Muller reveals an effort 569 to exaggerate the linear extrapolation range. Third, Muller 570 ( 1950b) makes an error in his statement that the linearity 571 was shown with a dose rate "less than 0.001 r per minute" 572 when the actual value was 0.00165 r/min. 573 Dispatch : 1-8-2013 0 LE 611 CP Pages : 19 0 TYPESET 611 DISK Arch Toxicol Table 1 Hermann J Mulier and Curt Stem quotes on low-dose linearity References Quote Muller ( l94X) Page 462 " ... the frequency of the mutations induced will be proportional to the total dose of radiation received over an unlimited period of time." "There is then absolutely no threshold dose, unlike what is true of many other biological effects of radiation, and even the most minute dose carries a definite chance of producing mutations-a chance exactly proportional to the size of that dose." Muller ( 1952) Page 317 "In making our calculations it is safe, as both the earlier (6-10) and the more recent (11-15) works have agreed, to accept the principle that the frequency of the gene mutations produced is simply (linearly) proportional to the amount of the total accumulated dose received, as expressed in r units. Moreover, as some of these same studies show, this relation holds within wide limits, regardless of how short and concentrated or dilute and protracted the exposure may have been, or whether it was given in one treatment or many." "There are good theoretical grounds for inferring that these principles hold true no matter how small the total dose, or the dose per unit time. Of course, such a sweeping conclusion necessarily involves an extrapolation from actual data. Not until recently has it been possible, because of technical difficulties, to test the mutagenic effectiveness of doses lower than about 13 r per day, totaling 400 r (11-13), and even the most recent work goes down no lower than about 2.5 r per day, totaling 25 r (14, 15)." Stem ( 1950) Page 433 "The proportionality rule has been proven to hold over a wide range. Figure 155 shows that, for Drosophila, the relation is essentially linear over the range from 25 r to several thousand r. It has further been shown that the frequency of induced mutations is independent of the time over which the radiation is applied." Stem ( 1960) Page 491 "It has been established for a variety of experimental organisms that the number of mutations induced by radiation is tional to the dose. This proportionality has been proven to hold over a wide range of dosages. Figure 202 shows that, for Drosophila, the relation is essentially linear over the range 25-12,500 r (insects, unlike mammals, can survive after exposure to many thousands of roentgens).

It would be desirable to extend the data toward dosages lower than 25 r, for instance, to 10 r, 5 r, and still lower. Since, however, the expected differences are small between the rate of mutations in not-artificially irradiated control organisms and that in organisms exposed to low artificial doses, it is difficult to obtain significant results even with large experiments." 574 Impact of the Stern and Muller deceptions 575 Effect on the radiation genetics literature/community 576 In the aftermath of his Nobel Prize Lecture, Muller pub-577 lished his Lecture in the Journal of Heredity in 1947 578 (Muller 19!7), assuring its broader distribution.

Within 579 4 months of the Noble Prize Lecture, he gave a lecture 580 to the New York Academy of Medicine during which he 581 affirmed his Nobel Prize Lecture message, stating that 582 there was "absolutely no threshold dose" for mutations 583 and that induced mutational response was proportional to 584 the total dose (Table I). This presentation was published in 585 the Academy's journal (Muller !948) soon thereafter.

Stem 586 (! 950) also cited Spencer and Stern ( 1948) and Uphoff and 587 Stem ( 1949) in his acclaimed textbook, emphasizing that 588 the dose response for mutations was linear (Table ! ). 589 These follow-up activities by Stern and Muller had an 590 impact on other leading radiation geneticists influencing 591 them to adopt the linearity dose-response interpretation.

592 Table 2 provides a series of quotations from subsequent 593 publications of leading contemporary radiation geneticists.

594 The quotes are numerous, varied, and a fair representa-595 tion of what each author stated. These comments strongly Journal : Large 204 Article No : 1105 MS Code : 1105 support the conclusion that there was a generally consist-596 ent view that the nature of the dose response in the low-597 dose zone for mutations was linear. Most of these quotes 598 directly cite the research of Stem and his colleagues as pro-599 viding the key evidence supporting linearity, especially that 600 of Spencer and Stern (I 948) and Uphoff and Stem ( 1949). 601 This demonstrates the significance and success of the Stem 602 mediated manipulation of the Caspari and Uphoff studies 603 in affecting mutation dose-response beliefs of key research 604 leaders of the radiation genetics community.

605 Effect on the BEAR I Committee/Genetics Panel 606 Crow (I 995) noted the following in his historical recount-607 ing of the BEAR I Committee Genetics Panel: "the debate 608 over the nature of the dose response for ionizing radiation 609 and mutations had been decided before the convening of the 610 BEAR Committee in November 1955." The accepted view 611 was clear and unified; the answer for the dose response 612 question for mutagenicity was "linearity at low dose." 613 When reading the transcripts of the BEAR I Committee 614 Genetics Panel, one is struck by the absence of debate and 615 even discussion on the issue of dose response (e.g., linear-616 ity vs. threshold).

To illustrate the fact that the decision on 617 Dispatch : 1-8-2013 0 LE li1l CP Pages : 19 0 TYPESET li1l DISK Arch Toxicol Table 2 Radiation genetics quotations about the mutation dose-response following Hermann J Muller's Nobel Prize and Curt Stem's (with Spencer, Caspari and Uphoff) mutagenicity papers References Catcheside ( 1950) Glucksmann ( 1950) Lefevre ( !950) Sax (J'JSO) Higgins (I 951) Stone (!952) Singleton ( 1954a) Kelner et al. ( !955)

Quotes Page 592 "The induced mutation is proportional to the total dose over the whole range investigated, down to total doses as small as 25 r. There is good reason to conclude that there is no threshold dose, i.e., no dose so small that it gives no tional effect. Also, the intensity of the radiation appears to be without effect on the frequency of mutation induced by a given total dose. A dose of 50 r given in a fraction of a minute appears to give no greater effect than the same dose given in the course of a few weeks. There is no threshold, no time factor, and no recovery, the effects being tive." Page 42 "The induction of gene mutations is linearly proportional to dose even down to levels of 25 r (Spencer and Stern l94X)." Page 341 "It has been amply verified that the number of mutations produced by X-rays is linearly proportional to the total dose applied, even when the total dose received is very small (see Spencer and Stern !9-tR). Further, the number of tions produced is independent of the rate of dosage (Uphoff and Stern 1949)." Page 332 "The early work by Muller and by Timofeeff-Ressovsky showed a linear relationship between X-ray dosage and tion frequency in Drosophila.

It was also found that the induced mutation rate was independent of radiation intensity.

From these observations it was concluded that the X-ray-induced mutations are produced by single 'hits,' and that there is no threshold effect. Spencer and Stern (2) found no increase over the spontaneous mutation rate by irradiating Drosophila for 21 days at 2.5 r/day, but later experiments by Uphoff and Stern (3) indicated that low intensities are effective." Page9 "As a result of exhaustive experiments on the genetics of the fruit fly, of mice and of many plants, it is held that the number of induced mutations bears a linear relationship to the total amount of radiation absorbed by the sensitive volume of the cell and is independent of either the duration or the intensity of exposure.

Consequently, a long exposure to low-level radiation would have the same genetic effect as shorter exposure to a higher level. Experiments of Spencer and Stern ( J 948) on the fruit fly show that the percentage of sperm containing a sex-linked lethal mutation is increased about .002 per r of radiation exposure and that 50 r exposure is required to double the natural mutation rate." "Spencer and Stern (I.e.) conclude their exhaustive study of the validity of the linear relationship between radiation exposure and mutation frequency with the statement (p. 64): ' ... for radiation with X-rays, dosages as low as 25 r produce mutations as drastic in their effects and in the same proportion to the dosage as do exposures to high dosages. If an extrapolation is permissible, one may assume that there exists no tolerance dose below which mutations are not induced." "The classical hit theory of induction of mutations, particularly the linear relation between dosage at low levels and mutation rate, has been questioned by Caspari and Stern (

who found no significant difference in mutation rates in the sperm of the fruit fly between controls and experimentals exposed to 2.5 r per day for 21 days. Uphoff and Stern ( l 949), however, after further tests, concluded that low-level radiation does produce mutations in fruit-fly sperm and that the apparent inconsistencies of previous results were due to different experimental techniques and errors in sampling." Page 657 "There is no threshold for genetic mutations

... " (cited Muller reference 1950, J Cell Comp Physiol 35(suppl I ):9-70.) Page 598 (Discussion) "That a non-linear relationship exists between dose rate of chronic gamma radiation and mutation rate of endosperm characters seems to have been well established by these experiments.

This was shown quite conclusively by portionately higher mutation rates at the higher dosages, and was definitely indicated by the fact that there seems to be a threshold of dosage required to raise the mutation rate from the spontaneous level to a detectable increase over that level." Page 599 "These data (i.e., data shown in Singleton 195*la study) showing a definite threshold are in contrast to the Drosophila data of Spencer and Stern ( 194S), where no threshold was indicated even when low doses of radiation were used. In their experiments the effects of acute radiation were studied. Caspari and Stern ( 19.fR), studying chronic gamma radiation, found no increase over the controls for doses of 2.5 r/day for 21 days. However, it was later demonstrated by Uphoff and Stern ( 1949) that the controls used by Caspari and Stern had an abnormally high sex linked lethal frequency and that actually there was an effect of the chronic gamma radiation of 2.5 r/day." Page 36 "The linear mutation-dose curve indicated for X-ray induced drosophila lethals (Lethals-Dros:X) is perhaps best exemplified by the data of Spencer and Stem (53) for sex linked Jethals and may be considered as the classical type of mutation-dose relation.

Interpreted within the target theory, the linear relation indicates that a single hit is sufficient to produce a mutation." Journal : Large 204 Anicle No : 1105 MS Code : 1105 Dispatch : 1-8-2013 D LE 0 CP Pages : 19 0 TYPESET 0 DISK 618 619 620 621 622 623 624 625 626 627 628 629 630 631 632 633 634 635 636 637 638 639 640 641 642 643 644 Arch Toxicol Table 2 continued References Quotes Nybom et a!. ( 19'i6) Page 81 "In this connection references may be made to the concordant results of Uphoff and Stem ( 1949) who did not find any threshold in Drosophila after low dose rates. A similar result was published by Sax (I 950) using chronic irradiation of Tradescantia pollen." Lewis (!9:'i7) Page 971 (columns 2 and 3) (This Science article was reprinted in Congressional Testimony) "Gene mutation has long been known to show a linear relationship with respect to dose of ionizing radiation from ies with Drosophila.

This linearity has been extended by Spencer and Stem (43) to doses of 50 and 25 roentgens.

Gene mutation is also known to be directly proportional to the accumulated dose of radiation, even when the radiation is chronically administered at a relatively low dose rate, as in the studies of Uphoff and Stem (44)." Norwood (19)8) Page 1929 "Several geneticists 4 have sketched the background which has lead to the concern of this study. Briefly, realization that radiation increases the mutation rate dates back 30 years to Muller's experiments with fruit flies4e. Spencer and Stem.5 using more than 50 million flies, showed that genetic damage was proportional to dosage in the important range of 25 to 50 r. Concern has been heightened by recent findings 41 that exposure of mice to a given quantity of radiation increases the mutation rate by about 15 times as much as does an equal exposure of Drosophila, which had formerly served as the sole basis for inferring*

human risks." Spear ( 195R) Page 20 "There is general agreement, however, that mutations can be produced with very low dosage down to a level which approaches natural background (Uphoff and Stern 19-W)." Newcombe ( 1960) Page 331 "One basic premise which has not so far been seriously challenged is that the number of gene mutations resulting from irradiation varies in direct proportion to the dose. In other words, there is no threshold level of radiation below which the mutations will not be produced." "In the fruitfly the curve has, by dint of considerable work, been pushed to within 25 roentgens of the origin (Caspari and Stem I '!48; Spencer and Stern 19*+11; Uphoff and Stem !949) (3, 4, 5)." LNT had already been settled prior to the creation of the BEAR I Committee, there was no discussion of the scien-tific foundations of the LNT, including any documenting of its theoretical basis and experimental support, including its strengths and limitations.

As noted above, the Genet-ics Panel placed a high priority on the chronic exposure experiments published under the leadership of Curt Stem. Yet these studies, even ignoring the control group problems of the Uphoff and Stem experiments, had little or no risk assessment relevance.

That is, these were sex-linked reces-sive lethality studies in which the spermatozoa were depos-ited in the spermatheca of the female. The females were then placed into a type of specialized experimental "hiber-nation" in which there was a profound alteration of the diet and a lowering of the temperature, changes designed to prevent egg production.

The females (with the deposited spermatozoa) were then exposed for 21 days (24 h/day) to gamma irradiation.

After the 21 days, the dietary and envi-ronmental conditions were changed to permit egg laying so that the testing for sex-linked recessive lethal mutations could take place. In effect, Stem exposed the spermatozoa to ionizing radiation for the equivalent of an entire lifespan, something comparable to a 70-80-year human lifespan.

The spermatozoa are known to be highly compromised, having lost much of their normal repair capability.

The study represented a worse case exposure scenario, that is, selection of a very susceptible developmental stage linked Journal : Large 204 Article No : 1105 MS Code : 1105 to a profoundly extended and highly unrealistic exposure period. In effect, the study was a chronic exposure to a cell type that has only a very short developmental stage. The basic concept of the study was not appropriate for a chronic exposure with risk assessment application.

The BEAR I Committee incorrectly accepted Stem and Muller's concept of "chronic" for risk assessment purposes as did the entire field and regulatory agencies.

While the BEAR I committee relied upon the findings of the Drosophila research directed by Curt Stem, it failed to cite other similarly large-scale Drosophila studies (Bon-nier and LUning 1949; Bonnier et a!. 1949) in which the lowest total dose was 8 r, below the lowest dose (25 r) of the Spencer and Stem ( l94R) findings.

These papers docu-mented the response of several single genetic loci (e.g., white and forked loci) to which their detailed statistical analysis for mutational studies was applied. The analysis revealed a linear dose response in the dose range of 700--2,800 r, whereas the linearity response was not observed in the low-dose range (8-16 r), where the data were sup-portive of a threshold response.

The authors also sug-gested that the difference in the shape of the dose response between high and low doses was indicative of differing dose-dependent mechanisms.

At the high doses, the lin-ear dose response was consistent with the target theory of Timofeeff-Ressovsky et a!. ( 1935), whereas at lower doses mutational effects could be due to the effects of chemical Dispatch : 1*8-2013 0 LE 0 CP Pages : 19 D TYPESET 0 DISK 645 646 647 648 649 650 651 652 653 654 655 656 657 658 659 660 661 662 663 664 665 666 667 668 669 670 671 672 mutagens (i.e., hydroxyl radicals from the hydrolysis of 673 water). The dose-dependent mechanism-based hypothesis 674 of Bonnier and colleagues (Bonnier and Liining !949; Bon-675 nier et al. 1949) was soon supported with experimental data 676 (Haas et al. 1950; MacKey 195 I; Liining 1954; Barron 677 1954). According to Barron (1954), "it is dangerous, how-678 ever, to extrapolate from experimental data with large doses 679 of radiations to what might take place with small doses. In 680 biological systems the effect of ionizing radiations differs 681 qualitatively when the radiation dose is changed. Small 682 doses act by indirect action and produce mainly oxidations.

683 Large doses act by two mechanisms," that is, free radical 684 formation via water hydrolysis and by a direct collision, 685 which is consistent with the target theory. 686 The Bonnier and Liining ( 1949) (Bonnier et al. 1949) 687 papers were also critical of the use of sex-linked recessive 688 lethal experiments for estimating responses in the low-dose 689 zone due to the "impossibility of differentiating between 690 true lethals and semi lethals, and the fact that there are sev-691 era! hundreds of targets per chromosome ready for lethal 692 mutations

... " The lack of target specificity would repre-693 sent an important limitation in the interpretation of dose-694 response relationships and their potential application to a 695 mechanism-based risk assessment process. Bonnier et al. 696 (1949) also provided a detailed statistical reanalysis of 697 the Spencer and Stern ( 1948) data challenging the broadly 698 accepted conclusion that the linearity response applied 699 across the entire dose-response range, including the lower 700 dose range. None of these fundamental technical issues 701 were discussed by the BEAR I committee.

702 Another relevant aspect of the discussion on the nature 703 of the mutation dose response involved the research 704 of Arnold H. Sparrow and W. Ralph Singleton of the 705 Brookhaven National Laboratory.

Chairman Warren 706 Weaver introduced their research and its relevance to the 707 BEAR I Committee/Genetics Panel (Weaver W., Febru-708 ary 5-6, 1956, see page 110-Transcript) (BEAR I 1956). 709 The discussion of the Sparrow and Singleton data was then 710 led by Committee member Berwind D. Kaufmann, who 711 claimed to have copied several tables from their paper. 712 He stated that Sparrow and Singleton showed that 0.41 r 713 per day yielded a modestly elevated (i.e., less than twice 714 the control values) but statistically significant effect on 715 micronuclei formation.

What Kaufmann failed to inform 716 the Committee was that Sparrow and Singleton ( !953) spe-717 cifically stated that a threshold response had been observed 718 at a lower dose. In fact, there was no discussion concern-719 ing their threshold dose-response statement by the BEAR 720 I Committee/Genetics Panel. The data in Table 2 (page 35) 721 of the published paper by Sparrow and Singleton ( 1953) 722 show that 0.084 r per day caused no significant increase in 723 micronuclei.

This recounted activity of the BEAR I Com-724 mittee/Genetics Panel demonstrates that it either ignored or J oumal : Large 204 Article No : 1105 MS Code : 1105 Arch Toxicol was misled on the published findings of Sparrow and Sin-725 gleton as the data did not support the pre-determined linear 726 dose-response conclusion.

This analysis also suggests that 727 the BEAR I Committee/Genetics Panel was very selec-728 tive in their choice of what data to consider and that such 729 decisions reveal a prevailing bias supportive of LNT model 730 acceptance.

731 Since 0.41 r per day of radiation in the Sparrow and Sin-732 gleton (1953) hypothesis study is more than 1,000 times 733 greater than the naturally occurring intensity, these data 734 do not support the theory that the spontaneously occurring 735 micronuclei are produced by naturally occurring ionizing 736 radiation.

The findings of Sparrow and Singleton

(! l)5l) 737 were similar to that of Giles (I ')40) from Harvard who 738 showed that when Tradescantia were "subjected to irradia-739 tion 1 ,000 times that due to natural radiation

.... no increase 740 in aberration was found." Other experiments by Giles indi-741 cated that even using ionizing radiation at some 1,800-fold 742 above background no impact on the occurrence of sponta-743 neous mutations occurred.

744 It is possible to obtain a sense of the personal views of a 745 number of the members of the BEAR I Committee/Genet-746 ics Panel on the matter of dose response via two contempo-747 rary publication avenues: Testimonies at a 1957 Congres-748 sional Hearings (Table 3) and journal publications in the 749 open literature (Table 4) such as a special issue of Scientific 750 American on ionizing radiation and several other journals.

751 Based on these collective comments, it follows that the 752 BEAR I Committee/Genetics Panel report and an article in 753 the journal Science (Table 5) summarizing the report of the 754 Genetics Panel were replete with statements asserting lin-755 earity at low dose. 756 Placing the new Muller and BEAR I Genetics Panel 757 developments in perspective 758 The story of Muller's Nobel Prize Lecture is important 759 for its history of science implications, as well as its role 760 in affecting the decision of the US National Academy of 761 Sciences (NAS) to recommend a linearity dose-response 762 policy for assessing risks to the genome from ionizing 763 radiation, replacing the threshold dose-response model. 764 This formal recommendation initiated a series of advi-765 sory and regulatory dominoes in essentially all countries 766 to adopt linearity and apply it to somatic effects, that is, 767 cancer risk assessment, for ionizing radiation and later for 768 chemical carcinogens (Calabrese 2009). The linearity deci-769 sion of the NAS BEAR I Committee/Genetics Panel was 770 strongly championed by Muller, the titular leader of radia-771 tion geneticists and with strong ties to all radiation geneti-772 cists on the BEAR I Committee/Genetics Panel. In fact, the 773 switch to linearity, which was ushered into the international 774 Dispatch : 1*8*2013 0 LE li1l CP Pages : 19 0 TYPESET li1l DISK Arch Toxicol Table 3 BEAR I Committee Genetics Panel member quotes at Joint Committee on Atomic Energy-1957 References Muller ( l 956) Crow ( l957a) Glass ( 19:'i7a) Muller ( 1957a) Muller ( 1957b) Joint Committee on Atomic Energy ( 1957) Quotes Page 392 "In material of varied kinds, but more especially in Drosophila, there is good evidence that over a considerable range of dose (in Drosophila, from some 50 r to more than I ,000 r, a more than 20-fold range) the frequency of point tions (like that of chromosome breaks) is directly proportional to dose." Page 1013 "4. Evidence from experimental animals, principally Drosophila, indicates that the number of mutations produced is strictly proportional to the amount of radiation received.

There are departures from this straight-line relationship at high doses, but these are too high to be likely to be encountered in any ordinary human situation.

It is technically impossible to test this relationship for the very lowest doses, but the straight-line relation holds down to the smallest amounts that have been studied." "For these reasons a simple proportionality between the amount of radiation and the number of mutations is fully accepted by geneticists." "The proportionality between dose and mutation production holds irrespective of the intensity or spacing of the dose." Representative Holifield (page 1013) questions Dr. Crow: "This, then, would establish as far as the majority of the geneticists are concerned the principle of linear progression in deleterious effects of radiation regardless of amount?" Dr. Crow answers: "That is correct. A nonthreshold situation, to put this in yesterday's vocabulary." "This means that there is no such thing as a safe dose of radiation to the population.

Any amount of radiation, however, small, that reaches the gonads-testes or ovaries-<>f a person who may later reproduce, involves a risk proportional to that amount." Page 1030 "The data are most extensive for the fruitfly and the lowest dose that has actually been studied is 25 r." Page 1031 "Because a mutation can be produced by a single ionization in the right place, there is no threshold below which the amount of radiation is too small to produce mutations-that is, every dose produces mutations with a probability equal to its magnitude." "This is to repeat what Dr. Crow said, that there is no safe dose of mutation.

This curve continues down without any threshold until it hits the zero point. .. " Page 1052 "In respect to the fact that probably there is no threshold, that these effects are proportional to the dose, in this respect these effects of radiation-and also the leukemia-<>n the exposed individual himself resemble those produced by the radiation in weakening descendants." "You have heard Dr. Glass and Dr. Crow say that geneticists are convinced that there is no threshold for the genetic effects and that others, too, now accept that principle for the genetic effects." "If this is true of these other effects, and it is certainly time we knew whether it was-I think the evidence is convincing that it is-then this important resemblance between the effects on later generations and on the exposed generation is probably not an accidental resemblance.

For there is growing reason to infer that this shortening of life and the other long delayed damage done to an exposed individual have their basis in damage done to the genetic material-the chromosomes and their contained genes-<>f the body's ordinary cells, those of the blood, skin, glands, and so forth, similar to the damage done in his reproductive cells that is passed on to later generations." Page 1056 "Through work on the fruitflies where we have the most exact knowledge to date, unless Dr. Russell has more exact knowledge on mice now, we can get a kind of minimum estimate of the amount of damage to the children by a given amount of irradiation of the parents." Page 1066 "Since there is much evidence indicating a linear relation between the radiation dose and the frequency of the induced point mutations, even at extremely low doses, and the exactly cumulative nature of these radiation effects, it becomes possible to arrive at probable estimates of the minimum damage done to subsequent generations by any given chronic or acute exposure of parents." Page 1067 " ... leukemia and some other malignancies, the induction of which may also be linearly dependent upon radiation dose ... " Page 12 " ... geneticists believe that the direct proportion applied down to zero dose-that is, that there exists no safe "threshold" below which the dose produces no damage, and that damage occurs from any irradiation of the genetic cells, no ter how small the dose." Journal : Large 204 Article No : 1105 MS Code : 1105 Dispatch : 1-8-2013 0 LE li1l CP Pages : 19 0 TYPESET li1l DISK 775 776 777 778 779 780 781 782 783 784 785 786 787 788 789 790 791 792 793 794 795 796 797 798 799 800 Arch Toxicol Table 4 BEAR I Committee Genetics Panel member quotes on low-dose linearity in journals after the BEAR I Committee References Quotes Crow ( 1957h) Page 19 (column 2) "2. The number of mutations produced is directly proportional to the dose in roentgens.

The linear propor-tionality over wide dose ranges has been shown in several organisms, especially in Drosophila." "Experimental verification in Drosophila has been carried to as low as 25 r ... " Page 20 (column I) "The proportionality between dose and mutation production holds irrespective of intensity or spacing, ... " Page 20 (column 2) "The conclusions of the previous section imply that there is no such thing as a "safe" dose. Any increase in radiation, however, small, involves a risk proportional to that amount." Glass (1957h) Page 956 "Our present evidence indicates that the frequency of these point mutations always increases linearly with the radiation dose (Fig. I). In Drosophila studies this holds over the range from 25 r to 6,000 r. In some plants, the linear range has been extended down to about 5 r. In mice, the linearity in relation to dose holds over the range from 300 r to 600 r, and there is no sign that it does not hold at lower doses. This linear proportionality to dose, over and above the spontaneous frequency of mutation, implies that (a) as long as dosage is measured in terms of roentgens, that is, in terms of the ionization produced by the radiation, absorbed quanta do not interact to produce effects, but are individually effective; and (b) there is no sign of a threshold dose below which mutations are not produced.

Rather, even the lowest doses are proportionally mutagenic, and all doses, however, distributed, are additive or cumulative in effect." Beadle ( 1959) Pages 225 and 226 " ... thus there is probably no threshold below which radiation will produce no mutations.

Since there is no repair mechanism, once the mutation process is complete, mutations induced at different times will tend to accumulate in a line of descent. .. " Hollaender and Stapleton (I 959) "In sum, cell studies have served to elucidate the basic mechanism by which ionizing radiation damages the living organism.

They have provided no evidence that there is a true threshold of dosage below which ionizing radiation produces no harmful effects ... " community by the BEAR I Committee Genetics Panel, is the most significant action in regulatory environmental public health history with ever expanding social, political, economic, and public health implications (Hamblin :2007). The present paper provides the first documentation of how Muiler (Muiier 1950a, b, l954a, b) himself used the carefuiiy constructed activities of Stem (described in detail in Calabrese 20 I I b) to enhance the concept of linearity and to protect his reputation.

Muiler lent credibility to the tech-nical note of Uphoff and Stem ( 1949) while further mar-ginalizing the Caspari and Stem study results (Caspari and Stem 1948). The stakes were high on multiple levels and these core individuals knew it. Stem and Muiier needed to prevent the acceptance of the Caspari and Stem (I 94R) study findings in order to sustain the single-hit linearity model. They also needed any criticisms of the Spencer and Stem (1948) and Uphoff and Stem ( 1949) papers to be muted. They were successful as other leaders of the radia-tion genetics community simply failed to address the seri-ous limitations of the Spencer and Uphoff findings while incorrectly asserting that the Cas pari and Stem ( 1948) paper suffered from an aberrantly high control value, sim-ply re-stating the demonstrably incorrect, but authoritative conclusion of Muiier ( J950a). Despite the fact that Caspari had successfuiiy rebutted the first challenge of Stem concerning the control group <fl Springer Journal : Large 204 Article No : 1105 MS Code : 1105 spontaneous mutation rate, there is no evidence that he disputed the control group mutation rate reversal decision of Stem barely a year later and of Muiler's equaiiy strange affirmation of Stem's position as weii (Muiier I950a, b). A January 27, 1949, Jetter from Cas pari to Stem supported the publication of the Uphoff and Stem ( 1949) paper now adopting part of the mantra of Stem, that is, that there is considerable variability in the mutagenic frequency of sperm prolongedly stored in the spermatheca.

This conclu-sion provided the opportunity to rehabilitate the inexplic-itly low control group values of Uphoff. Caspari, however, would not go so far as to also state that his control values were unusuaiiy high. At the time of the Uphoff and Stem ( 1949) article, there were only two papers published in the literature (Rajewski and Timofeeff-Ressovsky

!939; Kauf-mann 1947) on aged sperm and mutation and the published abstract of Muiier ( 1946b ). Each supported the mutation frequency of Caspari. These findings are consistent with subsequent mutation frequencies in aged sperm stored in the spermatheca of female Drosophila (Byers 1954; Byers and Muiier 1952; Rinehart 1969; Graf 1972; Muiier et a!. 1961 ). Muiler et a!. ( 1961) stated that "The data clearly showed a rise in mutation frequency (averaging some .06 percent of recessive lethals in the X chromosome per week) resulting from storage of the mature spermatozoa in the female" (page 213). Note the striking similarity of how Dispatch : 1-8-2013 0 LE li1l CP Pages : 19 D TYPESET li1l DISK 801 802 803 804 805 806 807 808 809 810 811 812 813 814 815 816 817 818 819 820 821 822 823 824 825 826 Arch Toxicol Table 5 Low-dose linearity quotation in the journal Science from article summarizing the findings of the BEAR I Committee Genetics Panel References Quotes BEAR I (1956) Page 1159 (column 2) " ... the genetic damage done, however, felt and, however, measured, is roughly proportional to the total mutation rate." Page 1160 (column I) "3) Any radiation dose, however, small, can induce some mutations.

There is no minimum amount of radiation dose, that is, which must be exceeded before any harmful mutations occur." Page 1160 (bottom column I) "The probable number of additional induced mutations occurring in an individual over a period of time is by and large proportional to the total dose of extra radiation received, over that period, by the reproductive organs where the germ cells are formed and stored." Page 1160 (top column 2) "The total dose of radiation is what counts, this statement being based on the fact that the genetic damage done by radiation is cumulative." Page 1162 (column 2)-how harmful are radiation-induced mutations? "I) Thus the first and unanimous reply to the question posed by the title to this section is simply this: Any radiation is genetically undesirable, since any radiation induces harmful mutations.

Further, all presently available scientific tion leads to the conclusion that the genetic harm is proportional to the total dose ... This tells us that a radiation dose of 2X must be presumed to be twice as harmful as a radiation dose of X ... " Page 1164 (column I) " ... for there is no such figure other than zero." [referring to whether there is an amount of radiation which is genetically harmless (preceding phase)) Page 1164 (column I) "As geneticists we say: keep the dose as low as you can." Page 1165 (last sentence) "From the point of view of genetics, they are all bad." (referring to the effect of exposures to ionizing radiation) 827 Uphoff and Stem (I 947) characterized Muller's data some 828 14 years earlier, "a weekly increase of about 0.07 % ... " 829 The 0.06 % increase would yield an estimated 0.28 % 830 (i.e., 0.06 % x 3 weeks + 0.10% background

= 0.28 %) 831 mutation incidence after 3 weeks, consistent with the Cas-832 pari and Stern ( 1948) findings, the logic used in Uphoff 833 and Stern ( l94 7) and with the Muller ( 1946b) statement 834 that "spermatozoa aged several weeks in the female may 835 contain several times as many mutations as they originally 836 had." Furthermore, the reported inter-study variability for 837 mutations of aged sperm and/or stored sperm aged in the 838 spermatheca appears modest with 95 % confidence inter-839 vals typically being about +/-25-30 % of the mean. The 840 attempt by Stern, therefore to assert that the very low val-841 ues of Uphoff reflected a highly variable response endpoint 842 was not supported in the contemporary and subsequent lit-843 erature. Stern never argued his case by a comparative data 844 assessment nor did he address the apparent contradiction 845 with the Muller data and comments which he (i.e., Stem) 846 previously used when he concluded that the Caspari data 847 were credible while those of Uphoff were not. He simply 848 made an authoritative declaration that was accepted without 849 question or comment by the radiation genetics community.

850 BEAR I Committee/Genetics Panel 851 The BEAR I Committee/Genetics Panel was comprised 852 of outstanding scientists and national leaders. Despite 853 their significant individual accomplishments in scientific Journal : Large 204 Article No : 1105 MS Code : 1105 and radiation genetics domains, the committee as a whole 854 lacked extensive experience in conducting low-dose, 855 dose-response studies. Only two of the members had 856 extensive direct experimental dose-response experience 857 (i.e., Demerec and Russell) up to the time of the BEAR I 858 meetings.

This experience was essential for evaluating the 859 nature of the dose response in the low-dose zone. Of these 860 two, Demerec had the most extensive and varied experi-861 ence having dealt with multiple models and agents as well 862 as different types of radiation.

His research experience 863 on dose response was spread over a 25-year period start-864 ing about 1931. Nonetheless, his dose-response experience 865 with Drosophila was limited to only a few high dose stud-866 ies during the 1930s, a key limitation.

Despite his signifi-867 cant and prolonged career at Oak Ridge, Russell was rela-868 tively new to the dose-response research area, with about 869 5-6 years experience at the start of the BEAR I Commit-870 tee in 1955. In the case of Russell, his developing research 871 findings with mice were still somewhat premature, having 872 little impact on BEAR I Committee/Genetics Panel con-873 elusions.

Among the remaining members of the commit-874 tee, Muller's principal dose-response experience is found 875 in the research of Hanson and Heys (! 929), and Oliver 876 ( 1930, 1931) at the University of Texas and Ray-Chaudhuri 877 ( 1944) at Edinburgh (completed in 1939), as well as his 878 consultant role with Stem from 1943 to 1946. Limited rei-879 evant low dose-response research based on the publication 880 record experience was found for Berwind Kaufmann.

Alex-881 ander Hollaender, PhD in physical chemistry, had made 882 Dispatch : 1-8-2013 0 LE li1l CP Pages : 19 0 TYPESET li1l DISK 883 important contributions on the effects of UV wavelengths 884 specificity on mutation in bacteria and fungi. He became 885 the director of radiation biology research at Oak Ridge, hir-886 ing Russell. Hollaender had no experience with Drosophila 887 research.

H. Bently Glass' low-dose experimental research 888 experience was limited during BEAR I, becoming far 889 more extensive only after BEAR I. Importantly, very lim-890 ited to no meaningful dose-response research experience is 891 apparent for the remaining 11 members [George W. Bea-892 die, Charles W. Cotterman, James F. Crow, Gioacchino 893 Failla, Clarence C. Little, James V. Nee!, Tracy M. Sonne-894 born, Alfred H. Sturtevant, Sewall Wright, Warren Weaver 895 (Chair), and Shields Warren] of the BEAR I Committee/

896 Genetics Panel. This situation resulted in the "senior" dose-897 response experience to reside with Demerec and Muller, 898 two individuals on record to save the "hit" model. 899 The geneticists on the BEAR I committee were princi-900 pally basic researchers; their experimental approaches were 901 neither dose response nor risk assessment oriented.

Even 902 Muller ( l 950a, h) claimed that the work of Spencer and 903 Uphoff (with Stern) at low doses would markedly extend 904 his and his students' (e.g., Hanson and Oliver) research 905 conducted at very high doses. Further, in the detailed com-906 ments that Muller sent to Stern about the Spencer (Lilly 907 Library ! 946, September

13) and Caspari (Lilly Library 908 ! 947a, January 14) manuscripts, nearly all dealt with fun-909 damental biological/genetic questions with little direct 910 relevance to risk asse.ssment.

Multiple study design issues 911 and other methodological/analysis problems documented in 912 Calabrese (20 1 I b) for the Spencer and Stern ( 1948) paper 913 were not identified by Muller (Lilly Library 1946, Septem-914 ber 13). The members of the BEAR I Committee/Genetics 915 Panel looked to Muller for leadership on matters related to 916 the dose-response.

However, Muller displayed critical lim-917 itations in assessing such studies based on his written state-918 ments. Thus, the methodological and analysis limitations of 919 the Spencer and Stern ( 1948) paper and the serious flaws 920 of the Uphoff and Stern (! 949) paper were missed by the 921 radiation genetics community and the BEAR I Committee/

922 Genetics Panel, a condition that continues (Lipshitz 2005). 923 Of further note is that Muller ( I946b) and Kaufmann 924 ( l 94 7) published findings on the control group mutation 925 rate of aged Drosophila sperm that supported the findings 926 of Caspari and Stern ( 1948). Kaufmann worked closely 927 with and under the direction of Demerec at Cold Spring 928 Harbor at that time. Furthermore, an October 7, 1947, letter 929 (i.e., 6 weeks before submitting his paper to Genetics) from 930 Caspari to Stern (American Philosophical Society 1947g, 931 October 7) stated that "I have discussed the paper (the 932 Caspari/Stern manuscript) with Demerec and Kaufmann.

933 Both did not find very much to suggest ...... Both Demerec 934 and Kaufmann were impressed by the amount of material 935 which we have. The ageing effect in our experiments is Journal : Large 204 Article No : 1105 MS Code : 1105 Arch Toxicol of the same order of magnitude as that found by Timofe-936 eff and Kaufmann." In fact, Caspari and Stern ( 1948) cited 937 a 1947 paper by Kaufmann as support for control group 938 values of their study. Muller and Kaufmann, both BEAR I 939 committee members, therefore, reported research on muta-940 tion incidence of Drosophila aged sperm findings con-941 sis tent with the findings of the Cas pari and Stern ( 1948) 942 paper. Thus, the BEAR I Committee/Genetics Panel should 943 have been informed on the issue of control group valid-944 ity by Demerec, Kaufmann, and/or Muller as it related to 945 the research of the Caspari and Uphoff studies. However, 946 based on the transcripts of the BEAR I Committee/Genet-947 ics Panel, Demerec, Kaufmann and Muller did not provide 948 this information.

Knowledge of the mutation rates in aged 949 Drosophila sperm should have led to a reconsideration of 950 the Caspari and Stern (I 948) paper as well as generated 951 serious questions about the findings and interpretations 952 of the Uphoff and Stern ( 1949) data. This was a key issue 953 affecting which study would be relied upon by the BEAR I 954 committee.

By their actions, the BEAR I committee Genet-955 ics Panel came to the erroneous conclusion that the Cas-956 pari study was unreliable due to its "unusually high control 957 group value." 958 The future of ionizing radiation risk assessment was 959 largely determined by the actions of a few, by the failure of 960 the scientific community, especially the radiation genetics 961 community, to probe deeper into the key findings of Stern 962 and his colleagues and journals such as Science that pub-963 lished influential but poorly documented findings (Uphoff 964 and Stern 1949). As has been pointed out, the linear-965 ity paper of Spencer and Stern ( 1948) was burdened with 966 numerous methodological limitations that only recently 967 have been documented, as well as statistical analysis limita-968 tions that challenged the conclusion of linearity at low dose 969 (Bonnier and LUning 19-f9; Bonnier et a!. 1949) while the 970 Cas pari and Stern ( 1948) findings supporting a threshold 971 perspective were unfairly marginalized (Calabrese 20 I I b). 972 Furthermore, the BEAR I Committee/Genetics Panel failed 973 to require Stern to provide the promised detailed account-974 ing for the Science article (Uphoff and Stern I 949) upon 975 which they so heavily relied. 976 According to Muller (J 950a, b), by 1950, the radiation 977 genetics community had accepted the linearity risk assess-978 ment paradigm (Table 2). Their belief was based largely 979 on the fruit-fly work of Stern and his associates as well as 980 the leadership, prestige, and authority of Muller, as few of 981 the geneticist members of the BEAR I Committee/Genet-982 ics Panel had relevant experience with low-dose research.

983 By the time, the National Academy of Sciences BEAR I 984 Committee/Genetics Panel convened, therefore, the deci-985 sion over the nature of the response in the low-dose zone 986 had been decided by the radiation genetics community 987 as there was no dispute or even debate within the BEAR 988 Dispatch : 1*8-2013 0 LE li2l CP Pages : 19 0 TYPESET li2l DISK Arch Toxicol 989 I Committee/Genetics Panel over the adoption of linearity 990 to replace the threshold model for germ-cell mutagenic-991 ity (Crow 1995). The actions of Stern and Muller had led 992 the way, assuring that the ends (i.e., linearity) justified the 993 means (i.e., unfair/improper scientific evaluation).

In fact, it 994 is from this heritage and upon this foundation that regula-995 tory cancer risk assessment theory and practice in the USA 996 and throughout the world was built. 997 Conclusions 998 1. This paper provides specific documentation of how 999 Hermann J. Muller supported and extended the like 1000 actions of Curt Stern to prevent the scientific com-1001 munity from discovering Muller's Nobel Prize lecture 1002 deception and to promote his ideological goal of line-1003 arity at low dose for ionizing radiation risk assessment 1004 (Table 6). 1005 2. Muller strengthened the questionable actions of Stern 1006 in key publications in early 1950s while improperly 1007 discrediting the threshold findings of Caspari and sup-porting the "uninterpretable" data of Uphoff to achieve 1008 a linearity interpretation.

The bases of these actions are 1009 documented in this paper. 1010 3. The paper shows how the actions of Stern and Muller 1011 affected numerous publications and the dose-response 1012 beliefs of leaders of the radiation genetic community 1013 and the NAS BEAR I Committee/Genetics Panel, 1014 affecting the adoption of linearity at low dose for ion-1015 izing radiation-induced mutation and eventually for 1016 carcinogen risk assessment for ionizing radiation and 1017 chemical carcinogens.

1018 4. The findings demonstrate that the adoption of the LNT 1019 model for risk assessment lacked a proper scientific 1020 foundation, yet was accepted by regulatory and public 1021 agencies worldwide.

1022 Unresolved issues 1023 1. Why didn't Stern publish the follow-up detailed paper 1024 containing the entire methodology for all the relevant 1025 data for the Uphoff three experiments?

1026 Table 6 A summary concerning Muller's actions that affected the discrediting of Caspari's findings and acceptance of the Uphoff and Stem conclusions A five-page detailed letter sent from Muller to Stern dated January 14, 1947, concerning scientific strengths and limitations of the Caspari and Stem manuscript provided no comment on the control group lethality data Muller was actively researching the area of spontaneous mutations in sex-linked recessive lethality studies using aged sperm stored in the matheca of female fruit flies. This was the research method of the Caspari and Stem paper. Muller had been doing extensive research on this topic since the early 1940s. He was a leading authority on the topic Muller provided his spontaneous control group data to Stem section) in order to address the concern that Stem expressed about the apparently high control group values of Caspari Based on the data of Muller, Uphoff and Stem ( l9*I7) determined that the average weekly spontaneous mutation rate in Drosophila sperm stored in the spermatheca of the female was about 0.07 %, yielding an additional mutation increase in about 0.21 %by 3 weeks, the length of the Caspari sperm storage time. The 0.21 %increase would be added to a background value of about 0.10 %, yielding an estimated control group value of about 0.31 %. The 95 %confidence intervals were about +/-0.07 %, with an approximate range of 0.24-0.38

%. The values were obtained when studies were conducted at about 25 °C. At the lower temperature of 18 oc used by Caspari, it was estimated by Stem (and Uphoff) that the rate of increase might be reduced to 0.05 % per week. This would result in an estimated value for the Caspari control of about 0.25 %, nearly identical to his final adjusted value (i.e., 0.2489%) Based on these data, Uphoff and Stem ( 19*17) concluded that the Muller data supported the Caspari conclusion that his control data were well within the normal range and not unusual or aberrant.

The Muller data lead Uphoff and Stem ( 1947) to conclude the Uphoff findings were uninterpretable Continued research in the area of spontaneous mutation in sperm stored in the spermatheca by Muller and his graduate students at the University of Indiana were consistent with this conclusion and quantitative assessment (Byers 1954; Byers and Muller Graf 1972). These findings were also consistent with that published by other researchers as well (Kaufmann I 9-1 7; Rinehart 1969) Based on this information, the statements of Muller that Caspari's control group data were unusually high are inconsistent with: (I) His own data and that published by other researchers; (2) his previously detailed assessment of the Caspari data; (3) how Uphoff and Stern (I <J-17) evaluated the Muller data, an evaluation that Muller was knowledgeable of, based on an acknowledgment in the Uphoff and Stem ( 19.:\7) paper, and (4) internal written correspondence between Stem and Caspari This assessment indicates that Muller's statements that Caspari's control group data were unusually high and adversely affected Caspari's old interpretation are contradicted by the body of evidence While Muller repeatedly challenged the credibility of the Caspari findings by attacking his control group data, he made no statement about the reliability of the extremely low control group data of Uphoff. In fact, he would consistently cite the Uphoff and Stem ( 1 ')-19) paper as being a critical reference to support a linearity perspective The collective findings on these matters indicate that Muller displayed compromised scientific judgment, having a significant impact on the scientific literature and national and international risk assessment policy that continues to the present Journal : Large 204 Article No : 1105 MS Code : 1105 Dispatch : 1-8-2013 0 LE 6"1 CP Pages : 19 0 TYPESET 6"1 DISK 1027 2. Why didn't the radiation geneticist community demand 1028 that Stern publish these findings?

1029 3. Why didn't Stern address the scientific basis, if any, 1030 of why he reversed his position on the Uphoff control 1031 group data? 1032 4. Why didn't Caspari challenge any of the multiple 1033 papers that claimed that the Caspari control group data 1034 were unusually/abnormally high or that their paper 1035 displayed "different techniques" or had "errors in sam-1036 piing" that accounted for their threshold-like findings?

1037 5. Why did Muller agree to let Uphoff and Stem ( 19-1-7) 1038 acknowledge the use of his aged sperm data that sup-1039 ported the Caspari control groups findings and then 1040 repeatedly claim that Caspari's control group values 1041 were unusually high, adversely affecting the credibility 1042 of this paper? 1043 Acknowledgments The research on the topic of hormesis has been 1044 supported by awards from the US Air Force and ExxonMobil Founda-1045 tion over a number of years. Sponsors had no involvement in study 1046 design, collection, analysis, interpretation, writing, and decision to 1047 submit. 1048 Conflict of interest The author declares that there is no conflict of 1049 interest.

1050 Appendix 1051 Stem-Muller temporal letter exchange concerning the 1052 aged-stored sperm control mutation rate (Source: Lilly 1053 Library, Stem-Muller correspondence) 1054 Curt Stem wrote a letter to Hermann J. Muller on January 1055 22, 1947 (American Philosophical Society 194 7a), inform-1056 ing him that "At the present time it looks as if our new con-1057 trol data (probably the results of the first 3 months of the 1058 first Uphoff experiment; note that her first month's reading 1059 was an especially low mutation rate of 0.005 %) for aged 1060 sperm are considerably below those of Caspari's." He then 1061 asked Muller to "send me your figures on rate of sex-linked 1062 lethal in sperm aged several weeks, (most desirably, if you 1063 have them, data on 3 weeks), in comparison to control data 1064 from non-aged sperm?" 1065 On February 3, 1947 (Lilly Library 19-l?b, February 3), 1066 Muller answered by stating that" .... sperm of males which 1067 are about a week old and have been copulating freely (as in 1068 Caspari's experiment) during that period have only about 1069 .07 or .08 % of lethal. Thus, the latter sperm, after 3 weeks, 1070 should contain something like .28 % of lethal." 1071 On July 23, 1947 (American Philosophical Society 1072 1947b), Stem writes Muller again stating that "I have mis-1073 laid your letter of some months ago (February 3, 1947, let-1074 ter) in which you gave me some details of your own on the Journal : Large 204 Article No : 1105 MS Code : 1105 Arch Toxicol mutation rate under various physiological conditions.

May 1075 I therefore ask you two questions and will you permit me to 1076 use your answers in a report which I am just preparing for 1077 the Manhattan Project? Obviously, full credit for it would 1078 be given. The questions are: (I) What is the spontaneous 1079 mutation rate in sperm derived from Canton-special males 1080 of from 3-to 6 days old? (2) What is the weekly increase in 1081 mutation rate of sperm from such males stored in females?" 1082 On August 4, 1947 (Lilly Library 19l7c), Muller 1083 responds "When sperm were stored in females, there was a 1084 weekly increase in the mutation frequency of about 0.07 %, 1085 on the average." On August 7, 1947 (American Philosophi-1086 cal Society l947c), Stern cabled Muller asking him the 1087 temperature used and on August 8, 1947 (American Philo-1088 sophical Society 1947d), Muller answered via cable indi-1089 eating "25 °C." A subsequent undated letter, but most likely 1090 prior to September 9, 1947 (American Philosophical Soci-1091 ety 19-+ 7.; ), Muller noted "A recalculation of my data gives 1092 the figure of 0.08 % instead of 0.07 % as the frequency 1093 of lethal accumulating in mature sperm per week." Since 1094 Uphoff and Stern ( 19-+ 7) did not include this correction in 1095 their report to the AEC it suggests that this undated letter 1096 was received after submittal of their report to the AEC. 1097 The control value therefore used by Uphoff and Stem 1098 ( 1947) of 0.07 % for the estimated mutation rate of the 1099 sperm stored in the spermatheca was based on the earlier 1100 letter correspondence-supplied estimates of Muller (Lilly 1101 Library l9+7b, c, February 3 and August 4) which Muller 1102 later clarified as being slightly in error. 1103 The Caspari and Uphoff studies used Drosophila mela-1104 nogaster fruit flies, breeding Canton-wild-type (S). males 1105 with Muller-5 females. Muller claimed (Lilly Library 1106 1947c, August 4) that he never conducted mutation experi-1107 ments with aged males of the Canton-wild-type stock. 1108 Muller stated that he had tested the aged sperm mutation 1109 frequency in "a number of different stocks (of Drosophila 1110 males) without finding any difference." The rate of increase 1111 on a weekly basis was said to be 0.07 % on average. This 1112 value of 0.07 % is believed to be prior to the correction 1113 to 0.08 %. This suggests that Muller did not observe sig-1114 nificant inter-stock variation in mutation rates of the stored 1115 sperm. 1116 Stern seems to have completed his Uphoff and Stern 1117 ( 19!7) paper for the Manhattan Project during August, 1118 1947. Stern knew that Uphoff's mean mutation fre-1119 quency was 0.1682 % (0.1365-0.2097

% ). This suggests 1120 a weekly mean increase in mutation rate of 0.0227 % 1121 (0.0122-0.0366

%), far lower than the 0.07 or 0.08 % 1122 mean weekly increase in Muller. When Stern wrote to 1123 Muller on September 9, 1947, he stated that for the Can-1124 ton-special stock " ... the weekly increase is considerably 1125 less than that found by you and others. It seems to be 1126 much more of the order of 0.03-0.05." This September 9, 1127 Dispatch : 1*8*2013 0 LE li1l CP Pages : 19 0 TYPESET li1l DISK Arch Toxicol 1128 1947, letter was written probably just after the submission 1129 of the Uphoff and Stem (I 947) paper to the AEC, and 1130 definitely before the submission of the Caspari and Stern 1131 ( 194S) paper for publication by Genetics (i.e., November 1132 25, 1947). Thus, the judgments of Uphoff and Stern that 1133 found that Uphoff's data were "uninterpretable" and that 1134 supported the reliability of the Caspari control data were 1135 made with the information provided by Muller during the 1136 summer of 1947. The apparent argument that Stern seems 1137 to be suggesting in his September 9, 1947, letter to Muller 1138 is that the Canton-wild-type stored sperm in the female 1139 may yield uniquely lower control mutation values. The 1140 argument is tenuous as the far higher weekly rate was 1141 consistently shown by multiple investigators, and with 1142 multiple Drosophila stocks, only being low in two Uphoff 1143 experiments.

In fact, significant inter-strain differences on 1144 the frequency of dominant lethal mutations as induced by 1145 radiation were not reported in various Drosophila strains, 1146 including the Canton-special wild-type strain (Demerec 1147 and Fano !944; Stromnaes 195! ). This suggestion by 1148 Stern was not included in the Uphoff and Stern ( 194 7) 1149 report. 1150 This letter exchange between Stern and Muller fails to 1151 provide support for the later statements of Muller that Cas-1152 pari's control group was unusually high. The Muller data 1153 and statements also do not provide support for the con-1154 elusion that the low Uphoff control data were in a normal 1155 range. None of this information was provided by Stern in 1156 his Science publication to permit the scientific community 1157 to better evaluate the Uphoff and Caspari control group 1158 data. 1159 References 1160 American Philosophical Society (1946) Muller letter to Stem. Stem 1161 Papers, Muller File-Box 16, 12 Nov 1162 American Philosophical Society (1947a) Stem letter to Muller, 22 Jan 1163 1947 1164 American Society (1947b) Stem letter to Muller, 23 1165 July 1947 1166 American Philosophical Society (1947c) Stem letter to Muller 1167 (cable), 7 Aug 1947 1168 American Philosophical Society (1947d) Muller letter to Stem, 8 Aug 1169 1947 1170 American Philosophical Society (1947e) Stem letter to Muller, 9 Sept 1171 1947 1172 American Philosophical Society (1947f) Caspari letter to Stem. Stem 1173 Papers, 25 Sept 1947 1174 American Philosophical Society (1947g) Caspari letter to Stem. Stem 1175 Papers, Box 21, 7 Oct 1176 American Philosophical Society (Undated)

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