Memories of a friend and colleague - Takashi Sugimura.

Takashi Sugimura, M.D., Honorary President of the National Cancer Center in Tokyo, and former President of The Japan Academy, is regarded by many as a pre-eminent contributor to the field of environmental genotoxicology. His pioneering spirit led to many key discoveries over a long and distinguished scientific career, including the first preclinical models for gastric cancer, identification of novel mutagens from cooked food, and the development of fundamental concepts in environmental chemical carcinogenesis. With his passing on September 6, 2020, many will reflect on the loss of an astute and engaging "Scientific Giant," who with warmth and good humor maintained lasting friendships both at home and abroad, beyond his many important scientific contributions.

The mutagenesis moonshot: The propitious beginnings of the environmental mutagenesis and genomics society.

A mutagenesis moonshot addressing the influence of the environment on our genetic wellbeing was launched just 2 months before astronauts landed on the moon. Its impetus included the discovery that X-rays (Muller HJ. [1927]: Science 64:84-87) and chemicals (Auerbach and Robson. [1946]: Nature 157:302) were germ-cell mutagens, the introduction of a growing number of untested chemicals into the environment after World War II, and an increasing awareness of the role of environmental pollution on human health. Due to mounting concern from influential scientists that germ-cell mutagens might be ubiquitous in the environment, Alexander Hollaender and colleagues founded in 1969 the Environmental Mutagen Society (EMS), now the Environmental Mutagenesis and Genomics Society (EMGS); Frits Sobels founded the European EMS in 1970. As Fred de Serres noted, such societies were necessary because protecting populations from environmental mutagens could not be addressed by existing scientific societies, and new multidisciplinary alliances were required to spearhead this movement. The nascent EMS gathered policy makers and scientists from government, industry, and academia who became advocates for laws requiring genetic toxicity testing of pesticides and drugs and helped implement those laws. They created an electronic database of the mutagenesis literature; established a peer-reviewed journal; promoted basic and applied research in DNA repair and mutagenesis; and established training programs that expanded the science worldwide. Despite these successes, one objective remains unfulfilled: identification of human germ-cell mutagens. After 50 years, the voyage continues, and a vibrant EMGS is needed to bring the mission to its intended target of protecting populations from genetic hazards. Environ. Mol. Mutagen. 61:8-24, 2020. © 2019 Wiley Periodicals, Inc.

Adventures with Bruce Ames and the Ames test.

Bruce Ames has had an enormous impact on human health by developing facile methods for the identification of mutagens. This research also provided important insights into the relationship between mutagenesis and carcinogenesis. Bruce is a highly innovative and creative individual who has followed his interests across disciplines into diverse fields of inquiry. The present author had the pleasure of spending a sabbatical in the Ames lab and utilized the Ames test in multiple aspects of his research. He describes both in this honorific to Bruce on the occasion of his 90th birthday.

Historical perspective on the development of the genetic toxicity test battery in the United States.

The currently used genetic toxicity testing battery (the Ames Salmonella test, the in vitro mammalian cell mouse lymphoma assay and/or the in vitro mammalian cell chromosome assay, and the rodent bone marrow chromosome aberration or micronucleus assay) had its origins in the early-to-mid 1970s. By the late 1970s, a large number of genetic tests had been proposed or recommended by the US-EPA for identifying germ cell mutagens and carcinogens. After a number of modifications that were primarily directed toward minimizing the number of tests used, the test battery reached its current state in the mid-1980s. This test battery, with some minor modifications in the timing or ordering of the tests is mandated by regulatory authorities worldwide. Although it would be intellectually satisfying to presume that this compendium of tests was developed and selected for regulatory screening based solely on scientific grounds, it was actually based on a combination of scientific data, theoretical considerations, chance, and advocacy, and not always in equal proportions. The evolution of the current genetic toxicity test battery, and some of the activities and considerations that directed this evolution are described.

Low dose intoxication and a crisis of regulatory models. Chemical mutagens in the Deutsche Forschungsgemeinschaft (DFG), 1963-1973.

Regulation and the prevention of danger are among the main characteristics of the modern state. However, the idea and the conceptualization of what danger is have changed over time. The genealogy of these changes shows that the history of social change and the history of knowledge are well connected. The 1970s marked the start of a social transformation of Western industrialized societies. This article proposes that this transformation was connected with basic epistemic reconfigurations and that the genealogy of risk played a significant role. This thesis is explored through the example of DFG advisory politics. Beginning in the 1960s, the DFG expert commissions that had been established to make policy and regulation recommendations began to focus more and more on the health effects of environmental pollution. The Commission for Questions of Mutagenicity played a particularly interesting role because its recommendations resulted in the foundation of a research institution run by the DFG, the Central Laboratory for Mutagenicity Testing (CML). The challenges faced by the CML in mutagenic research and testing effected a crisis of the expert-based advisory politics of the Mutagenicity Commission and a fundamental shift in the way scientific (regulatory) knowledge was perceived and valued politically. The pattern of this crisis calls to mind the constellation of the "risk society", but as will be shown, the (re)balancing of science and politics/society presented here is more adequately understood within the framework of political epistemology.

Isolation of plasmid pKM101 in the Stocker laboratory.

pKM101 is a mutagenesis-enhancing resistance transfer plasmid (R plasmid) that was introduced into several tester strains used in the Salmonella/microsome mutation assay (Ames test). Plasmid pKM101 has contributed substantially to the effectiveness of the Ames assay, which is used on a world-wide basis to detect mutagens and is required by many government regulatory agencies for approval to market new drugs and other chemical agents. Widely used since 1975, the Ames test is still regarded as one of the most sensitive genetic toxicity assays and a useful short-term test for predicting carcinogenicity in animals. Plasmid pKM101, which is a deletion derivative of plasmid R46 (also referred to as R-Brighton after its origin of isolation in Brighton, England), has also been used to elucidate molecular mechanisms of mutagenesis. It was isolated in the laboratory of Professor Bruce A.D. Stocker at Stanford University as part of my doctoral research with 20 R plasmids. Professor Stocker's phenomenal insight into the genetics of Salmonella typhimurium and plasmid behavior was a major factor that led to the isolation of pKM101. This paper includes a tribute to Bruce Stocker, together with a summary of my research with mutagenesis-enhancing R plasmids and a brief discussion of the molecular mechanisms involved in pKM101 plasmid-mediated bacterial mutagenesis.

History and rationale of genetic toxicity testing: an impersonal, and sometimes personal, view.

Genetic toxicity testing is a necessary and pivotal component of product development and registration. This article traces the historical development and evolution of genetic toxicity testing, and the rationale for such testing, and identifies some of the individuals who played key roles in this process. The evolution of the present test batteries and some of the research and rationales behind the decisions to accept or reject tests are described.

History of the science of mutagenesis from a personal perspective.

A career in the study of mutagenesis spanning 50 years is a gift few scientists have been bestowed. My tenure in the field started in 1953, the year the structure of DNA became known (Watson and Crick [1953]: Nature 171:737). Before that time, it was suspected that DNA was the genetic material based on the research of Oswald T. Avery (Avery et al. [1944]: J Exp Med 79:137), but many scientists still believed that proteins or polysaccharides could be the genetic material. The present article describes a lifetime of personal experience in the field of chemical mutagenesis. The methods used to treat viruses with chemical mutagens were well developed in the 1950s. Here I review the early use of nitrous acid and hydroxylamine as mutagens in eukaryotes, the development of methods for the metabolic activation of mutagens by microsomal preparations, and the selection of a mutant tester set for the qualitative characterization of the mutagenic activity of chemicals. These studies provided critical background information that was used by Bruce Ames in the development of his Salmonella/microsome assay, widely known as the Ames test (Ames et al. [1973]: Proc Nat Acad Sci USA 70:2281-2285). This article also describes how a set of diagnostic chemical mutagens was selected and used to identify the molecular nature of gene mutations. Today, DNA sequencing has replaced the use of diagnostic mutagens, but studies of this kind formed the foundation of modern mutation research. They also helped set the stage for the organization of the Environmental Mutagen Society and the Environmental Mutagen Information Center, which are described. The article ends with the development of mammalian single-cell mutation assays, the first system for studying in vivo mutagenesis using recoverable vectors in transgenic animals, other mutation assays in intact mammals, and my thoughts on the critically important area of germ cell mutagenesis. This narrative is not a complete autobiographical account, in that I have selected only those experiences that I feel are important for the history of the field and the edification of today's students. I hope I have shown that science not only is a valuable pursuit but can also be fun, stimulating, and satisfying. A good sense of humor and the knowledge that many discoveries come by serendipity are essential.

Observations at the interface of mutation research and regulatory policy.

Between 1970 and 1975 developments in environmental mutagenesis proceeded with amazing speed. These developments were both structural and conceptual in nature. A new infrastructure was built and new concepts about how best to protect consumers from exposures to mutagens emerged. The internal dynamics within the Food and Drug Administration played an important role and is discussed with regard to modifications in testing protocols as well as changes in the overall approach used to protect consumers. It is clear that this exciting period in the early days of environmental mutagenesis has provided a base for growth and development of the field and continues to affect and guide future developments.

The origins of the back-mutation assay method: a personal recollection.

The back-mutation assay method for determining the mutagenicity of various treatments was first developed a little over 50 years ago and has been in continuous use ever since. Shortly after the method was first used it became evident that certain factors of cell density, composition of media, etc., had to be carefully controlled to preserve an acceptable reliability of the method. A factor of particular importance was the suppression of growth of back-mutant prototrophic cells by the large number of auxotrophic cells present, a phenomenon which later became known as the "Grigg Effect." This review describes the origins of the back-mutation method and of the confounding competitive suppression phenomenon, the cause of competitive suppression, methods of diagnosing whether it is likely to bias the interpretation of a particular back-mutation experiment, and an experimental design which removes it entirely as a possible source of error. A number of other phenomena, such as phenotypic lag and coincident mutation associated with back-mutation, are also discussed as possible sources of error.

Carcinogenicity and mutagenicity testing, then and now.

Cancer is a dread disease worldwide. Mortality of individuals suffering from cancer is high, despite the current improved methods of precocious detection, surgery and therapy. Prevention of cancer is the recognized goal of many activities in cancer research. This aim was recognized early to involve the bioassay of environmental chemicals or mixtures. The first such study involved application of coal tar to the ear of rabbits, and later on to the skin of mice. Subsequently, laboratory rats were introduced, and hamsters were utilized as a substitute for the unwieldy tests in rabbits. Investigators also became concerned with the mechanisms of carcinogenesis, and more definitive approaches to carcinogen bioassay in laboratory animals, as possible indicators of cancer risk in humans. These tests were expensive and lengthy, and did not serve the important purpose of accurately measuring risk of cancer to humans. Once it was realized that DNA and the genetic apparatus might be a key target, rapid bioassays in bacterial and mammalian cell systems were introduced successfully. Thus, batteries of tests are now available to detect effectively human cancer risks, and provide novel approaches to determine the underlying mechanisms, as a sound basis for cancer prevention.